U.S. patent application number 16/346100 was filed with the patent office on 2019-08-22 for intestinal health promoting compositions.
This patent application is currently assigned to NSE Products, Inc.. The applicant listed for this patent is NSE Products, Inc.. Invention is credited to Shelly Hester, Angela Mastaloudis, Steven M. Wood.
Application Number | 20190255134 16/346100 |
Document ID | / |
Family ID | 62023893 |
Filed Date | 2019-08-22 |
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United States Patent
Application |
20190255134 |
Kind Code |
A1 |
Mastaloudis; Angela ; et
al. |
August 22, 2019 |
INTESTINAL HEALTH PROMOTING COMPOSITIONS
Abstract
Compositions for promoting intestinal health are disclosed and
described. In one example, the composition can include a
combination of cyanidins and delphinidins, in an amount sufficient
to treat intestinal hyperpermeability. In a further example, the
composition can further comprise a prebiotic blend and
fructooligosaccharides. Further presented herein, is a method of
treating a condition or disorder related to gastrointestinal health
in a subject. In one example, the method can include maximizing
tight junction integrity in epithelial cells of gastrointestinal
tract of the subject.
Inventors: |
Mastaloudis; Angela;
(Holladay, UT) ; Wood; Steven M.; (Santaquin,
UT) ; Hester; Shelly; (Draper, UT) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
NSE Products, Inc. |
Provo |
UT |
US |
|
|
Assignee: |
NSE Products, Inc.
Provo
UT
|
Family ID: |
62023893 |
Appl. No.: |
16/346100 |
Filed: |
October 27, 2016 |
PCT Filed: |
October 27, 2016 |
PCT NO: |
PCT/US2016/059226 |
371 Date: |
April 29, 2019 |
Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61K 31/352 20130101;
A61K 9/14 20130101; A61P 3/00 20180101; A61P 5/50 20180101; A23V
2250/284 20130101; A61K 31/702 20130101; A61P 1/04 20180101; A61K
36/185 20130101; A61K 36/45 20130101; A61K 45/06 20130101; A61P
35/00 20180101; A61K 31/733 20130101; A23L 33/105 20160801; A61P
9/00 20180101; A61P 11/06 20180101; A61P 29/00 20180101; A61P 3/10
20180101; A61P 7/00 20180101; A61P 37/08 20180101; A61P 3/04
20180101; A23V 2250/2106 20130101; A61P 19/02 20180101; A23V
2002/00 20130101; A23V 2250/2104 20130101; A61P 31/04 20180101;
A61P 43/00 20180101; A23V 2250/5062 20130101; A61K 36/899 20130101;
A61P 31/00 20180101; A61P 1/14 20180101; A23P 10/40 20160801; A61P
25/28 20180101; A61P 1/00 20180101; A23V 2200/3202 20130101; A61K
31/352 20130101; A61K 2300/00 20130101; A61K 31/733 20130101; A61K
2300/00 20130101; A61K 31/702 20130101; A61K 2300/00 20130101; A61K
36/185 20130101; A61K 2300/00 20130101; A61K 36/45 20130101; A61K
2300/00 20130101; A61K 36/899 20130101; A61K 2300/00 20130101 |
International
Class: |
A61K 36/45 20060101
A61K036/45; A61K 36/899 20060101 A61K036/899; A61K 31/352 20060101
A61K031/352; A61K 31/702 20060101 A61K031/702; A61K 31/733 20060101
A61K031/733; A61K 9/14 20060101 A61K009/14; A61P 1/00 20060101
A61P001/00; A23L 33/105 20060101 A23L033/105 |
Goverment Interests
GOVERNMENT INTEREST
[0001] A portion of the research for this invention was made with
government support under a grant awarded by the National Institute
of Food and Agriculture. The government has certain rights in the
invention.
Claims
1. An intestinal health promoting composition, comprising a
combination of cyanidins and delphinidins, in an amount sufficient
to treat intestinal hyperpermeability.
2. The composition of claim 1, wherein the cyanidins and the
delphinidins are collectively present in an amount that maintains
intestinal permeability.
3. The composition of claim 1, wherein the cyanidins and the
delphinidins are collectively present in an amount that reduces
intestinal hyperpermeability.
4. The composition of claim 1, wherein a source of at least one of
the cyanidins and the delphinidins are derived from a black rice
component, a blueberry component, a black current component, a
crowberry component, a bilberry component, black chokeberry
component, or a combination thereof.
5. The composition of claim 4, wherein the source of the cyanidins
and the delphinidins are derived from the black rice component, the
blueberry component, and the black current component.
6. The composition of claim 4, wherein the composition comprises
the black rice component and the black rice component is derived
from a member selected from the group consisting of black rice
kernels, black rice concentrate, black rice extract, black rice
powder, or a combination thereof.
7. The composition of claim 6, wherein the black rice component is
a black rice extract.
8. The composition of claim 7, wherein the black rice extract is
derived from black rice kernels.
9. The composition of claim 6, wherein the black rice component
comprises from about 2.5 wt % to about 20 wt % of the active
fraction of the composition.
10. The composition of claim 6, wherein the black rice component
comprises from about 10 wt % to about 15 wt % of the active
fraction of the composition.
11. The composition of claim 6, wherein the black rice component
comprises from about 2.5 wt % to about 7.5 wt % of the active
fraction of the composition.
12. The composition of claim 6, wherein the black rice component
has a standardized anthocyanin content ranging from about 10 wt %
to about 30 wt %.
13. The composition of claim 6, wherein the black rice component
has a standardized anthocyanin content of about 20 wt %.
14. The composition of claim 6, wherein the black rice component
has a standardized anthocyanin content of about 25 wt %.
15. The composition of claim 6, wherein the black rice component is
derived from Oryza sativa L.
16. The composition of claim 4, wherein the composition comprise
the blueberry component and the blueberry component comprises a
member selected from the group consisting of blueberry fruit,
blueberry extract, blueberry concentrate, blueberry juice,
blueberry powder, or a combination thereof.
17. The composition of claim 16, wherein the blueberry component is
a blueberry powder.
18. The composition of claim 16, wherein the blueberry component is
a blueberry juice.
19. The composition of claim 16, wherein the blueberry component
comprises from about 1 wt % to about 30 wt % of the active fraction
of the composition.
20. The composition of claim 16, wherein the blueberry component
comprises from about 1 wt % to about 10 wt % of the active fraction
of the composition.
21. The composition of claim 16, wherein the blueberry component
comprises from about 25 wt % to about 30 wt % of the active
fraction of the composition.
22. The composition of claim 16, wherein the blueberry component
has a standardized anthocyanin content ranging from about 0.5 wt %
to about 30 wt %.
23. The composition of claim 16, wherein the blueberry component
has a standardized anthocyanin content ranging from about 0.5 wt %
to about 5 wt %.
24. The composition of claim 16, wherein the blueberry component
has a standardized anthocyanin content ranging from about 20 wt %
to about 30 wt %.
25. The composition of claim 16, wherein the blueberry component is
derived from Vaccinium uliginosum L.
26. The composition of claim 4, wherein the composition comprises
the black current component and the black current component
comprises a member selected from the group consisting of black
current fruit, black current extract, black current concentrate,
black current juice, black current powder, or a combination
thereof.
27. The composition of claim 26, wherein the black current
component is a black current extract.
28. The composition of claim 26, wherein the black current
component comprises from about 0.5 wt % to about 15 wt % of the
active fraction.
29. The composition of claim 26, wherein the black current
component comprises from about 1 wt % to about 5 wt % of the active
fraction.
30. The composition of claim 26, wherein the black current
component has a standardized anthocyanin content ranging from about
20 wt % to about 40 wt %.
31. The composition of claim 26, wherein the black current
component has a standard anthocyanin content of about 30 wt %.
32. The composition of claim 26, wherein the black current
component is derived from Ribes nigrum.
33. The composition of claim 4, wherein the composition comprises
the crowberry component and the crowberry component comprises a
member selected from the group consisting of crowberry fruit,
crowberry extract, crowberry concentrate, crowberry juice,
crowberry powder, or a combination thereof.
34. The composition of claim 33, wherein the crowberry component
comprises crowberry fruit.
35. The composition of claim 33, wherein crowberry component
comprises crowberry extract.
36. The composition of claim 33, wherein the crowberry component
comprises from about 1 wt % to about 30 wt % of the active fraction
of the composition.
37. The composition of claim 33, wherein the crowberry component
comprises from about 5 wt % to 25 wt % of the composition.
38. The composition of claim 33, wherein the crowberry component
has a standard anthocyanin content ranging from about 40 wt % to
about 50 wt %.
39. The composition of claim 33, wherein the crowberry component
has a standardized anthocyanin content of about 46.7 wt %.
40. The composition of claim 28, wherein the crowberry component is
derived from Empetrum nigrum.
41. The composition of claim 4, wherein the composition comprises
the bilberry component and the bilberry component comprises a
member selected from the group consisting of bilberry fruit,
bilberry extract, bilberry concentrate, bilberry juice, bilberry
powder, or a combination thereof.
42. The composition of claim 41, wherein the bilberry component
comprises bilberry extract.
43. The composition of claim 41, wherein the bilberry component
ranges from about 0.5 wt % to about 30 wt % of the active fraction
of the composition.
44. The composition of claim 41, wherein the bilberry component
ranges from about 2 wt % to about 20 wt % of the composition.
45. The composition of claim 41, wherein the bilberry component has
a standardized anthocyanin content ranging from about 1 wt % to
about 30 wt %.
46. The composition of claim 41, wherein the bilberry component has
a standardized anthocyanin content ranging from about 5 wt % to
about 15 wt %.
47. The composition of claim 41, wherein the bilberry component
comprises 36 wt % anthocyanins as measured by HPLC or 25 wt %
anthocyanins as measured by UV.
48. The composition of claim 41, wherein the bilberry component is
derived from Vaccinium myrtillus.
49. The composition of claim 4, wherein the source of at least one
of the cyanidins and the delphinidins is derived from a black rice
component, a blueberry component, and a black current
component.
50. The composition of claim 49, wherein the black rice component,
the blueberry component, and the black current component have a
ratio of about 1:1:1.
51. The composition of claim 49, wherein the black rice component,
the blueberry component, and the black current component have a
ratio of about 1:1.4:4.3.
52. The composition of claim 1, further comprising a prebiotic
blend.
53. The composition of claim 52, wherein the prebiotic blend
comprises inulin.
54. The composition of claim 53, wherein the inulin is a chicory
inulin having oligosaccharides and polysaccharides with fructose
units linked by .beta. (2-1) linkages.
55. The composition of claim 53, wherein the inulin is derived from
banana, onion, flour, garlic, asparagus, wheat, rye, leeks,
chickory root, sugar beets, or a combination thereof.
56. The composition of claim 53, wherein the inulin comprises from
about 15 wt % to about 60 wt % of the composition.
57. The composition of claim 53, wherein the inulin comprises from
about 15 wt % to about 25 wt % of the composition.
58. The composition of claim 53, wherein the inulin comprises from
about 40 wt % to about 60 wt % of the composition.
59. The composition of claim 52, wherein prebiotic blend comprises
fructooligosaccharides.
60. The composition of claim 59, wherein the fructooligosaccharides
are short chain fructooligosaccharides (DP.ltoreq.5).
61. The composition of claim 60, wherein the short chain
fructooligosaccharides are derived from sucrose.
62. The composition of claim 60, wherein the short chain
fructooligosaccharides are derived from sugarcane.
63. The composition of claim 59, wherein the fructooligosaccharides
comprise from about 10 wt % to about 40 wt % of the active fraction
of the composition.
64. The composition of claim 59, wherein the fructooligosaccharides
comprise from about 10 wt % to about 20 wt % of the active fraction
of the composition.
65. The composition of claim 59, wherein the fructooligosaccharides
comprise from about 25 wt % to about 40 wt % of the active fraction
of the composition.
66. The composition of claim 59, wherein the fructooligosaccharides
comprise galactooligosaccharides.
67. The composition of claim 1, further comprising a prebiotic
blend of inulin and fructooligosacharides.
68. The composition of claim 67, wherein the inulin and
fructooligosacharides collectively comprise from about 55 wt % to
about 95 wt % of the composition.
69. The composition of claim 67, wherein a combine source of the
cyanidins and the delphinidins comprise from about 5 wt % to about
50 wt % of the composition.
70. The composition of claim 1, further comprising a
pharmaceutically acceptable carrier.
71. The composition of claim 1, further comprising a sweetener, a
preservative, a flavoring, a thickener, or a combination
thereof.
72. The composition of claim 1, wherein the composition is an oral
dosage form.
73. The composition of claim 72, wherein the oral dosage form
comprises a capsule, a tablet, a powder, a beverage, a syrup, a
gummy, a wafer, a confectionery, a suspension, or a food.
74. The composition of claim 72, wherein the oral dosage form
comprises a powder.
75. The composition of claim 72, wherein the oral dosage form is
designed to be administered to a subject in need thereof once per
day.
76. The composition of claim 75, wherein the oral dosage form is
designed to be administered to the subject at morning.
77. The composition of claim 72, wherein the oral dosage form
comprises a member selected from the group consisting of a black
rice component, a blueberry component, a black current component, a
crowberry component, a bilberry component, black chokeberry
component, or a combination thereof.
78. The composition of claim 77, wherein the oral dosage form
comprises the black rice component and the black rice component
ranges from about 500 mg to about 800 mg of the oral dosage
form.
79. The composition of claim 78, wherein the black rice component
has a standardized anthocyanin content of about 15 wt % to about 30
wt %.
80. The composition of claim 77, wherein the oral dosage form
comprises the blueberry component and the blueberry component
comprises from about 100 mg to about 3,000 mg of the oral dosage
form.
81. The composition of claim 80, wherein the blueberry component
has a standardized anthocyanin content of about 0.5 wt % to about
25 wt %.
82. The composition of claim 77, wherein the oral dosage form
comprises the black current component and the black current
component comprises from about 200 mg to about 3,000 mg of the oral
dosage form.
83. The composition of claim 82, wherein the black current
component has a standardized anthocyanin content of about 2.5 wt %
to about 30 wt %.
84. The composition of claim 77, wherein the oral dosage form
comprises the crowberry component and the crowberry component
comprises from about 100 mg to about 1,000 mg of the oral dosage
form.
85. The composition of claim 84, wherein the crowberry component
has a standardized anthocyanin content of about 1 wt % to about 30
wt %.
86. The composition of claim 77, wherein the oral dosage form
comprises the bilberry component and the bilberry component ranges
from about 100 mg to about 700 mg of the oral dosage form.
87. The composition of claim 86, wherein the bilberry component has
a standardized anthocyanin content of about 30 wt % to about 40 wt
%.
88. The composition of claim 77, wherein the oral dosage form
comprises the black chokeberry component and the black chokeberry
component ranges from about 50 mg to about 700 mg of the oral
composition.
89. The composition of claim 88, wherein the black chokeberry
component has a standardized anthocyanin content of about 1 wt % to
about 35 wt %.
90. The composition of claim 77, wherein the oral dosage form
further comprises a prebiotic blend.
91. The composition of claim 90, wherein the prebiotic blend
comprise from about 1 grams to 2 grams of the oral dosage form.
92. The composition of claim 91, wherein the prebiotic blend
provides from about 1 gram to about 2 grams of fiber in the oral
dosage form.
93. The composition of claim 77, wherein the oral dosage form
further comprises a fructooligosaccharide.
94. The composition of claim 93, wherein the fructooligosaccharides
comprise from about 1 gram to about 1. 5 grams of the oral dosage
form.
95. The composition of claim 93, wherein the fructooligosaccharides
comprise from about 3 grams to about 4 grams of the oral dosage
form.
96. The composition of claim 72, wherein the oral dosage form
comprises from about 200 mg to about 300 mg of anthocyanins.
97. A method of treating a condition or disorder related to
gastrointestinal health in a subject comprising maximizing tight
junction integrity in epithelial cells of gastrointestinal tract of
the subject.
98. The method of claim 97, wherein the gastrointestinal health of
the subject is improved.
99. The method of claim 98, wherein improved gastrointestinal
health of the subject comprises improvements in the subject's bowel
habits.
100. The method of claim 98, wherein improved gastrointestinal
health of the subject comprises reducing bloating, discomfort, gas,
or a combination thereof in the subject.
101. The method of claim 98, wherein improved gastrointestinal
health of the subject comprises reducing intestinal
hyperpermeability.
102. The method of claim 98, wherein improved gastrointestinal
health of the subject comprises a reduction in intestinal
dysbiosis.
103. The method of claim 98, wherein the improved gastrointestinal
health of the subject comprises reduced calprotectin fecal
levels.
104. The method of claim 98, wherein the improved gastrointestinal
health of the subject comprises an increase short chain fatty acid
levels.
105. The method of claim 97, wherein the condition or disorder
comprises inflammation, inflammatory bowel disease, irritable bowel
syndrome, chronic intestinal diseases, celiac disease, Crohn's
disease, ulcerative colitis, food intolerances, dyspepsia, low
levels of chronic intestinal inflammation, gastrointestinal
infections, or a combination thereof.
106. The method of claim 105, wherein the inflammation is reduced
overall.
107. The method of claim 106, wherein the inflammation is reduced
by up to 73%.
108. The method of claim 75, wherein supplementation for 3 weeks
results in a modest reduction in inflammatory biomarkers.
109. The method of claim 97, wherein the condition or disorder
comprises insufficient absorption of nutrients, endotoxemia,
intestinal hyperpermeability, or a combination thereof.
110. The method of claim 97, wherein the condition or disorder
comprises obesity, obesity associated pathologies, allergies,
cardiovascular conditions, type I diabetes, type II diabetes,
rheumatoid arthritis, insulin resistance, cancer, metabolic
syndrome, asthma, neurodegenerative diseases, or a combination
thereof.
111. The method of claim 97, wherein the condition or disorder is a
cardiovascular condition.
112. The method of claim 111, wherein the cardiovascular condition
is an increase in high-density lipoprotein cholesterol for subjects
that are not taking cardiovascular medications.
113. The method of claim 111, wherein the cardiovascular condition
comprises a decrease in HbA1c levels.
114. The method of claim 113, wherein the decrease in HbA1c levels
is from pre-diabetic levels to normal levels.
115. The method of claim 111, wherein the cardiovascular condition
comprises a decrease in plasma zonulin levels.
116. The method of claim 97, wherein the condition is peripheral
insulin resistance and the method reduces peripheral insulin
resistance.
117. The method of claim 97, wherein condition or disorder is type
I diabetes or type II diabetes.
118. The method of claim 97, wherein the condition or disorder
comprises a nitric oxide related disorder, expression of iNOS,
expression of COX-2, NADPH oxidase, or a combination thereof.
119. The method of claim 97, wherein the condition or disorder
stems from pathogens, antigens, and pro-inflammatory factors that
pass through the tight junctions in the epithelial cells of the
gastrointestinal tract.
120. The method of claim 97, wherein maximizing tight junction
integrity comprises protecting the gastrointestinal tract of the
subject from TNF.alpha. induced permealization of a monolayer of
the epithelial cells.
121. The method of claim 120, wherein an amount of the protecting
is concentration dependent on an amount of cyanidins and
delphinidins in the subject's gastrointestinal tract.
122. The method of claim 97, wherein the method further comprises
increasing transepithelial electrical resistance in the epithelial
cells.
123. The method of claim 97, wherein the method further comprises
increasing FITC dextran paracellular transport.
124. The method of claim 97, wherein the condition stems from
pro-inflammatory factors and the pro-inflammatory factors comprise
advanced glycation end products.
125. The method of claim 97, wherein the condition stems from
pro-inflammatory factors and the pro-inflammatory factors comprise
lipopolysaccharides.
126. The method of claim 97, wherein the condition stems from
pro-inflammatory factors and the pro-inflammatory factors comprise
cytokines tumor necrosis alpha (TNF-.alpha.), IL-6, or a
combination thereof.
127. The method of claim 97, wherein the condition or disorder
relates to conditions or disorders associated with signaling
pathways NF-kB, ERK1/2, or a combination thereof.
128. The method of claim 97, wherein the maximizing of the tight
junction integrity mitigates high fat induced intestinal
permeabilization.
129. The method of claim 97, wherein the epithelial cells comprise
Caco-2 cell monolayers.
130. The method of claim 97, wherein the method further comprises
optimizing a balance of gut microbiota in the gastrointestinal
tract.
131. The method of claim 130, wherein optimizing the balance of gut
microbiota comprises increasing commensal bacteria levels in the
gastrointestinal tract.
132. The method of claim 131, wherein the commensal bacteria belong
to bifidobacteria genus.
133. The method of claim 131, wherein the commensal bacteria belong
to bacteroidetes phylum.
134. The method of claim 131, wherein the commensal bacteria
comprise bacterdies caccae, bacteriodes uniformis, or a combination
thereof.
135. The method of claim 131, wherein the increase in the commensal
bacteria after 8 weeks of daily administering the method to the
subject in need thereof was at least 20%.
136. The method of claim 130, wherein optimizing the balance of gut
microbiota comprises increasing diversity of bacteria.
137. The method of claim 136, wherein the diversity of bacteria
comprises at least 200 strains.
138. The method of claim 130, wherein the method further comprising
decreasing harmful gut bacteria.
139. The method of claim 138, wherein the harmful gut bacteria
comprise firmicutes.
140. The method of claim 139, wherein the decreasing of the
firmicutes after 8 weeks of daily administering the method to the
subject was greater than a 15% reduction.
141. The method of claim 140, wherein a ratio of
firmicutes:bacteriodetes decreased approximately 3% after 8 weeks
of administering the method to the subject.
142. The method of claim 138, wherein the harmful gut bacteria
comprise Actinobacteria.
143. The method of claim 142, wherein the decreasing of the
Actinobacteria after 8 weeks of daily administering the method to
the subject was at least 5%.
144. The method of claim 138, wherein the harmful gut bacteria
comprise Helicobacter pylori.
145. The method of claim 138, wherein the harmful gut bacteria
comprise Clostridium.
146. The method of claim 138, wherein the harmful gut bacteria
comprise Klebisella.
147. The method of claim 97, wherein the method comprises providing
a fuel source for commensal bacteria.
Description
BACKGROUND
[0002] The gastrointestinal system is a complex network of tissues,
organs, host cells, and bacterial cells. Aging, diets, medications,
disruptions in the intestinal microbiota, and an individual's
lifestyle conditions can negatively affect this network. These
effects can include disrupting the balance of the gut microbiota,
increased inflammation in the gastrointestinal tissues, and a
disruption in the epithelial cell barrier of the gastrointestinal
system. These effects can ultimately lead to systemic issues.
Formulation and methods that promote and restore intestinal health
would provide a benefit.
BRIEF DESCRIPTION OF THE DRAWINGS
[0003] Features and advantages of the invention will be apparent
from the detailed description that follows, and which taken in
conjunction with the accompanying figures together illustrate
features of the invention. It is understood that the figures merely
depict exemplary embodiments and results and are therefore, not to
be considered limiting in scope.
[0004] FIG. 1 schematically displays the human gastrointestinal
tract and related structures;
[0005] FIG. 2 schematically displays the small intestine and the
epithelial cell layer lining the intestines;
[0006] FIG. 3 schematically displays an overview of the tight
junctions in epithelial cells and their systemic impact when the
tight junction barrier is disrupted;
[0007] FIG. 4 schematically displays the intestinal epithelial cell
layer, tight junctions, the paracellular pathway and the
transcellular pathway;
[0008] FIG. 5 schematically displays the chemical structure of
cyanidin;
[0009] FIG. 6 schematically displays the chemical structure of
delphinidin;
[0010] FIG. 7 schematically displays the chemical structure of
petunidin;
[0011] FIG. 8 schematically displays the chemical structure of
peonidin;
[0012] FIG. 9 schematically displays the chemical structure of
malvidin;
[0013] FIG. 10 graphically displays the transepithelial electrical
resistance (TEER) method in accordance with one example of the
disclosure;
[0014] FIG. 11 graphically displays the TEER resistance for various
extracts in accordance with one example of the disclosure;
[0015] FIG. 12 graphically displays paracellular transport for
various extracts in accordance with one example of the
disclosure;
[0016] FIG. 13 graphically displays TEER resistance for various
extracts in accordance with one example of the disclosure;
[0017] FIG. 14 graphically displays PCP FITC-dextran for various
extracts in accordance with one example of the disclosure;
[0018] FIG. 15 graphically displays the TEER resistance for
cyanidins in accordance with one example of the disclosure;
[0019] FIG. 16 graphically displays the TEER resistance for
delphinidin in accordance with one example of the disclosure;
[0020] FIGS. 17 & 18 graphically display TEER resistance for
epicatechin in accordance with one example of the disclosure;
[0021] FIG. 19 graphically displays TEER resistance for catechin in
accordance with one example of the disclosure;
[0022] FIG. 20 graphically displays TEER resistance for total
catechins in accordance with one example of the disclosure;
[0023] FIG. 21 graphically displays average colon length of mice on
different diets in accordance with one example presented
herein;
[0024] FIG. 22 graphically displays average colon weight of mice on
different diets in accordance with one example presented
herein;
[0025] FIG. 23 graphically displays average colon weight/length of
mice on different diets in accordance with one example presented
herein;
[0026] FIG. 24 graphically displays body weight gain and FITC
dextran permeability of mice on different diets in accordance with
on example presented herein;
[0027] FIG. 25 graphically displays the FITC-DX paracellular
transport of mice on different diets in accordance with one example
presented herein;
[0028] FIG. 26 graphically displays endotoxin levels measured of
mice on different diets in accordance with one example presented
herein;
[0029] FIG. 27 graphically displays GTT (AUC) levels measured of
mice on different diets in accordance with one example presented
herein;
[0030] FIG. 28 graphically displays ITT (AUC) levels measured of
mice on different diets in accordance with one example presented
herein;
[0031] FIG. 29 graphically displays endotoxin and glucose tolerance
test levels measured in accordance with one example present
herein;
[0032] FIG. 30 graphically displays endotoxin and fasting insulin
levels measured in accordance with one example presented
herein;
[0033] FIG. 31 graphically displays endotoxin and IL-6 test levels
measured in accordance with one example present herein;
[0034] FIG. 32 graphically displays endotoxin and IL-1.beta. levels
measured in accordance with one example presented herein;
[0035] FIG. 33 graphically displays endotoxin and IL-la test levels
measured in accordance with one example present herein;
[0036] FIG. 34 graphically displays HOMA-IR levels measured of mice
on different diets in accordance with one example presented
herein;
[0037] FIG. 35 graphically displays adiponectin levels measured of
mice on different diets in accordance with one example presented
herein;
[0038] FIG. 36 graphically displays leptin levels measured of mice
on different diets in accordance with one example presented
herein;
[0039] FIGS. 37 & 38 graphically displays triglyceride levels
measured of mice on different diets in accordance with one example
presented herein;
[0040] FIGS. 39 & 40 graphically displays cholesterol levels
measured of mice on different diets in accordance with one example
presented herein;
[0041] FIG. 41 graphically displays liver triglyceride levels
measured of mice on different diets in accordance with one example
presented herein;
[0042] FIG. 42 photographically displays mice livers extracted from
mice fed different diets prior to be euthanized in accordance with
one example presented herein;
[0043] FIG. 43 photographically displays mice droppings from mice
on different diets in accordance with one example presented
herein;
[0044] FIG. 44 graphically displays the average
firmicutes:bacterodetes ratio of the gut microbiome following
supplementation with a composition as presented herein;
[0045] FIG. 45 graphically displays calprotectin concentrations
following supplementation with a composition as presented herein
in;
[0046] FIG. 46 graphically displays a change in baseline BSS scores
and bowel movements following supplementation with a composition as
presented herein; and
[0047] FIG. 47 graphically displays the change in baseline scores
for bloating, abdominal pain, and gas following supplementation
with a composition as presented herein.
DETAILED DESCRIPTION
[0048] Before invention embodiments are disclosed and described, it
is to be understood that no limitation to the particular
structures, process steps, or materials disclosed herein is
intended, but also includes equivalents thereof as would be
recognized by those ordinarily skilled in the relevant arts. It
should also be understood that terminology employed herein is used
to describe particular examples only and is not intended to be
limiting. Unless defined otherwise, all technical and scientific
terms used herein have the same meaning as commonly understood by
one of ordinary skill in the art to which this disclosure belongs.
In the specification and the appended claims, the singular forms
include plural referents unless the context clearly dictates
otherwise.
[0049] As used herein, the singular forms "a," "an," and "the"
specifically provide express support for plural referents, unless
the content clearly dictates otherwise. For example, "a prebiotic
fiber" provides support for one or more prebiotic fibers.
[0050] The term "about" is used herein refers to a degree of
deviation. It means approximately, in the region of, roughly, or
around. When the term "about" is used in conjunction with a
numerical range, it modifies that range by extending the boundaries
slightly above and slightly below the numerical values set forth.
It is understood that support in this specification for numerical
values used in connection with the term "about" is also provided
for the exact numerical value itself as though "about" were not
used.
[0051] Concentrations, amounts, and other numerical data may be
expressed or presented herein in a range format. It is to be
understood that such a range format is used merely for convenience
and brevity and thus should be interpreted flexibly to include not
only the numerical values explicitly recited as the limits or
endpoints of the range, but also to include all the individual
numerical values and/or sub-ranges encompassed within that range as
if each numerical value (including fractions) and sub-range is
explicitly recited. As an illustration, a numerical range of "about
1 to about 5" should be interpreted to include not only the
explicitly recited values of about 1 to about 5, but also include
individual values and sub-ranges within the indicated range. Thus,
included in this numerical range for example, are individual values
such as 2, 2.6, 3, 3.8, and 4 and sub-ranges such as from 1-3, from
2-4, and from 3-5, etc., as well as 1, 2, 3, 4, and 5,
individually.
[0052] As used herein a "concentrate" refers to an extract of a
source that contains at least the same amount of active fractions,
compounds, or other constituents, in a smaller volume than in the
source itself. In one example, a "concentrate" may be a dried
powder derived from a component that does not include the use of
any solvents during the concentration process.
[0053] Comparative terms such as "more effectively," "greater
than," "improved," "enhanced," and like terms can be used to state
a result achieved or property present in a formulation or process
that has a measurably better or more positive outcome than the
thing to which comparison is made. In some instances comparison may
be made to the prior art or to the status of a property before
administration of a formulation or method that results in the more
positive outcome.
[0054] As used herein, "comprises," "comprising," "containing," and
"having" and the like can have the meaning ascribed to them in U.S.
patent law and can mean "includes," "including," and the like, and
are generally interpreted to be open ended terms. The terms
"consisting of" or "consists of" are closed terms, and include only
the components, structures, steps, or the like specifically listed
in conjunction with such terms, as well as that which is in
accordance with U.S. patent law. "Consisting essentially of" or
"consists essentially of" have the meaning generally ascribed to
them by U.S. patent law. In particular, such terms are generally
closed terms, with the exception of allowing inclusion of
additional items, materials, components, steps, or elements, that
do not materially affect the basic and novel characteristics or
function of the item(s) used in connection therewith. For example,
trace elements present in a composition, but not affecting the
composition's nature or characteristics would be permissible if
present under the "consisting essentially of" language, even though
not expressly recited in a list of items following such
terminology. When using an open ended term in the written
description, like "comprising" or "including," it is understood
that direct support should be afforded also to "consisting
essentially of" language, as well as, "consisting of" language as
if stated explicitly and vice versa.
[0055] The term "dosage unit" is understood to mean a single unit
of composition which is capable of being administered to a subject
or patient, to provide an amount of an active agent sufficient to
achieve, or contribute to, a therapeutic effect to be achieved. In
some embodiments, a "dosage unit" is a unit and that may be readily
handled and packed, remaining as a physically and chemically stable
unit dose comprising either the active ingredient as such or a
mixture of it with solid or liquid pharmaceutical vehicle
materials. Moreover, "dosage" and "dose" can refer to such a dosage
unit. Alternatively, a "dosage" or "dose" can encompass multiple
dosage units which collectively provide a desired amount of active
agent to be administered to a subject at a singular point in time.
Multiple doses or dosages can be utilized according to a schedule
in order to establish a dosing regimen.
[0056] The phrase "effective amount," "therapeutically effective
amount," or "therapeutically effective rate(s)" of an active
ingredient refers to a non-toxic, but sufficient amount or delivery
rates of the active ingredient, to achieve therapeutic results in
treating a disease or condition for which the active agent is being
delivered. It is understood that various biological factors may
affect the ability of a substance to perform its intended task.
Therefore, an "effective amount," "therapeutically effective
amount," or "therapeutically effective rate(s)" may be dependent in
some instances on such biological factors. Further, while the
achievement of therapeutic effects may be measured by a physician
or other qualified medical personnel using evaluations known in the
art, it is recognized that individual variation and response to
treatments may make the achievement of therapeutic effects a
subjective decision. The determination of a therapeutically
effective amount or delivery rate is well within the ordinary skill
in the art of pharmaceutical sciences and medicine.
[0057] The term "extract" refers to those substances prepared using
a solvent, e.g., ethanol, water, steam, superheated water,
methanol, hexane, chloroform liquid, liquid CO.sub.2, liquid
N.sub.2, propane, supercritical CO.sub.2, or any combination
thereof. Extracts, as used herein, can refer to an extract in a
liquid form, or can refer to a product obtained from further
processing of the liquid form, such as a dried powder or other
solid form. Extracts may take many forms including but not limited
to: solid, liquid, particulate, chopped, distillate, etc. and may
be performed by any number of procedures or protocols, such as
chopping, grinding, pulverizing, boiling, steaming, soaking,
steeping, infusing, applying a gas, etc., and may employ any
suitable reagents, such as water, alcohol, steam, or other organic
materials. Extracts typically have a given purity percentage and
can be relatively to highly pure. In some embodiments, extracts can
be phytoextracts made from specific parts of a source, such as the
skin, pulp, leaves, flowers, fruits, kernels, seeds, of a plant
etc., or can be made from the whole source. In some aspects, an
extract may include one or more active fractions or active agents.
In some aspects, the purity of an extract can be controlled by, or
be a function of the extraction process or protocol.
[0058] As used herein, "formulation" and "composition" can be used
interchangeably and refer to a combination of at least two
ingredients. In some embodiments, at least one ingredient may be an
active agent or otherwise have properties that exert physiologic
activity when administered to a subject.
[0059] Formulation or compositional ingredients included or recited
herein are to be presumed to be in wt % unless specifically stated
otherwise. In addition, ingredient amounts presented in the form of
ratios are to be presumed to be in wt % (e.g. % w/w) ratios.
[0060] The phrase, "intestinal hypermeability" refers to a higher
than normal (i.e. higher than that of an average subject)
permeability of the gastrointestinal system and can in some
instances refer to increased permeability in the stomach, small
intestines and/or large intestines.
[0061] As used herein, "linear inhibitory effect" or
"dose-response" refers to a linear decrease in secretion or
biosynthesis resulting from all concentrations of the inhibiting
material over a dose-response curve. For example, inhibition at low
concentrations followed by a failure of inhibition or increased
secretion at higher concentrations represents a lack of a linear
inhibitory effect.
[0062] The term "or" is used in the "inclusive" sense of "and/or"
and not the "exclusive" sense of "either/or."
[0063] As used herein, "pharmaceutically acceptable" refers
generally to materials, which are suitable for administration to a
subject in connection with an active agent or ingredient. For
example, a "pharmaceutically acceptable carrier" can be any
substance or material that can be suitably combined with an active
agent to provide a composition or formulation suitable for
administration to a subject. Excipients, diluents, and other
ingredients used in or used to prepare a formulation or composition
for administration to a subject can be used with such term.
[0064] The term "prevent" and its variants refer to prophylaxis
against a particular undesirable physiological condition. The
prophylaxis may be partial or complete. Partial prophylaxis may
result in the delayed onset of a physiological condition. An
individual skilled in the art will recognize the desirability of
delaying onset of a physiological condition, and will know to
administer the compositions of the invention to subjects who are at
risk for certain physiological conditions in order to delay the
onset of those conditions. For example, the person skilled in the
art will recognize that obese subjects are at elevated risk for
coronary artery disease. Thus, the person skilled in the art will
administer compositions of the invention in order to improve the
gut microbiota in an obese individual.
[0065] As used herein, a "subject" refers to an individual
receiving treatment. In one aspect, a subject can be a mammal. In
another aspect, a subject can be a human. In another aspect, the
subject can be a domesticated animal or livestock.
[0066] As used herein, "substantial," or "substantially," when used
in reference to a quantity or amount of a material, an effect, or a
specific characteristic of a composition thereof, refers to an
amount that is sufficient to provide an effect that the material or
a characteristic the material was intended to provide. The exact
degree of deviation allowable may in some cases depend on the
specific context. Similarly, "substantially free of" or the like
refers to the lack of an identified element or agent in a
composition. Particularly, elements that are identified as being
"substantially free of" are either completely absent from the
composition, or are included only in amounts which are small enough
so as to have no measurable effect on the composition.
[0067] As used herein, the term "insignificant" or "clinically
insignificant" refers to the extent of an effect of administering a
composition to a subject. For example, if the extent of the result
does not cause a clinical change in the subject, then the result is
clinically insignificant.
[0068] The terms "treat," "treating," or "treatment" as used herein
and as well understood in the art, mean an approach for obtaining
beneficial or desired results, including without limitation
clinical results in a subject being treated. Beneficial or desired
results can include, but are not limited to, alleviation or
amelioration of one or more signs or symptoms of a condition,
diminishment of extent of disease, stabilizing (i.e. not worsening)
the state of a disease or condition, delaying or slowing of disease
progression, amelioration or palliation of the disease state,
diminishment of the reoccurrence of disease, and remission (whether
partial or total), whether detectable or undetectable. "Treat,"
"treating" and "treatment" can also mean prolonging survival as
compared to expected survival if not receiving treatment and can be
prophylactic. Such prophylactic treatment can also be referred to
as prevention or prophylaxis of a disease or condition. The
prophylaxis may be partial or complete. Partial prophylaxis may
result in the delayed onset of a physiological condition.
[0069] As used herein, the term "solvent" refers to a liquid of
gaseous, aqueous, or organic nature possessing the necessary
characteristics to extract solid material from a plant product.
Examples of solvents would include, but not limited to, water,
steam, superheated water, methanol, ethanol, ethyl acetate, hexane,
chloroform, liquid CO.sub.2, liquid N.sub.2, propane, or any
combinations of such materials.
[0070] As used herein, a plurality of items, structural elements,
compositional elements, and/or materials may be presented in a
common list for convenience. However, these lists should be
construed as though each member of the list is individually
identified as a separate and unique member. Thus, no individual
member of such list should be construed as a de facto equivalent of
any other member of the same list solely based on their
presentation in a common group without indications to the
contrary.
[0071] Any steps recited in any method or process claims may be
executed in any order and are not limited to the order presented in
the claims unless otherwise stated.
[0072] The gastrointestinal system is a tract of connected and
related structures that are involved in the digestion of food and
absorption of energy and nutrients. The gastrointestinal tract,
shown in FIG. 1, consists of all the structures between the mouth
and the anus. The whole gastrointestinal tract is about nine meters
long and can be divided into an upper gastrointestinal tract and a
lower gastrointestinal tract. The lower gastrointestinal tract
includes the small intestine and the large intestine. The small
intestine is about 20 feet in length and has a highly folded
structure that includes finger like protections called villi.
Within the villi is a single layer of epithelial cells and a layer
of capillaries. The main function of the small intestine is to
absorb products of digestion. Nutrients can pass through the layer
of epithelial cells and enter the capillaries underneath. These
nutrients can eventually enter larger blood vessels and travel to
the liver where they are processed and regulated for release into
the body. The large intestine is about 3 feet long and is used to
collect the solid material that was not digested in the small
intestine and to absorb water. The large intestine is loaded with
bacteria that synthesize vitamins.
[0073] As previously mentioned, the small intestine has a barrier
composed of a single layer of epithelial cells. These epithelial
cells are sealed by tight junctions. The tight junctions can
modulate intestinal permeability by regulating paracellular
transport of water and ions and the transcellular pathway. See,
FIG. 3. In addition to being responsible for absorption of
nutrients, this cell layer also plays a role in maintaining the
mucosal immune homeostasis, preventing inflammation, and
constitutes the first line of defense against the entry of noxious
bacteria/bacterial toxins and/or other antigens that can initiate
chronic inflammation. Any bacteria, bacterial toxins, antigens,
water, and ions that pass through this cell layer can enter the
blood stream, affect other organs, and can have systemic effects on
the individual overall. See, FIG. 4. Lifestyle and dietary factors
such as high-intensity exercise, high-fat diets, and over nutrition
can also influence intestinal permeability and can play a role in
increasing toxin permeability across tight junctions and into
circulation.
[0074] Disruption of the tight junction results in a leaky tight
junction barrier and can lead to an increase in intestinal
permeability. Increased permeability can be a major factor in the
pathophysiology of several inflammation and obesity related
pathologies. Low levels of chronic inflammation can negatively
affect intestinal barrier permeabilization.
[0075] Tumor necrosis factor alpha (TNF.alpha.) can also be a
central underlying mediator. TNF.alpha. triggers apoptosis;
however, these changes in distribution occur through its capacity
to promote barrier permeabilization and expression of select tight
junction proteins. Ultimately, TNF.alpha. can play a role in
promoting tight junction barrier dysfunction. Loss of tight
junction functionality and increased intestinal permeability can be
contributors to the pathology of allergies (e.g. celiac disease),
inflammatory bowel diseases (Crohn's disease and ulcerative
colitis), food intolerances, dyspepsia, low levels of chronic
intestinal inflammation (e.g. those associated with obesity and
type I and II diabetes), insulin resistance, autism, multiple
sclerosis, malnutrition, metabolic syndrome, cancer, asthma, atopy,
and rheumatoid arthritis.
[0076] Another factor in the health of an individual's
gastrointestinal system is the microbiota of the gut. The
microbiota is a complex network of bacteria, archae, viruses, and
eukarya that influence an individual's health and physiology. The
gastrointestinal microbiota has been estimated to reach
approximately 3.9.times.10.sup.13 microorganisms. The composition
of an individual's microbiota can vary greatly in numbers,
diversity, histology, and activities. The richness and diversity of
the gut microbiota has been shown to be significantly influenced by
antibiotics, age, diet, ethnicity, geographical location,
physiological stress, psychological stress, and sex.
[0077] Alterations in the gut microbiota can cause an increase in
endotoxin production. A microbiota that has few and less diverse
beneficial bacteria and has greater numbers and more diverse
non-beneficial bacteria can occur with age and/or conditions
associated with accelerated aging, such as, obesity or a high fat
diet. An imbalance of the microbiota can also lead to increased
inflammation of the gastrointestinal lining, changes in the
integrity of the intestinal cell wall, and can led to gut
permeability. These changes can contribute to the incidence of
gastrointestinal infections, asthma/atopy, obesity, metabolic
syndrome, cancer, rheumatoid arthritis, Crohn's disease, and
ulcerative colitis. In contrast, a balanced and diverse microbiota
can provide resistance to infections, allow for healthy aging,
prevent intestinal disorders, contribute to polyphenol metabolism,
and can generate absorbable bioactives. The gut microbiome can also
play a role in metabolism, immune development, endocrine signaling,
and neurologic signaling.
[0078] Flavonoids can play an important role in the prevention and
amelioration of intestinal barrier permeabilization. The microbiota
in the gut can metabolobize flavoinoids releasing absorbable
bioactives. These bioactives can restore maintain and/or restore
the nutrient balance. Accordingly, flavonoids have the capacity to
inhibit inflammation, modulate select signaling cascades, and
regulate cellular redox status. However, the large number of
existing flavonoids with different chemical/spatial structures, and
the multiple metabolites generated at the large intestine by the
microbiota, rule out the possibility of simple generalizing the
health benefits and mechanisms of action of flavonoids as a
class.
[0079] Anthocyanins (AC) constitute one of the major flavonoid
subgroups, which are present and provide color to many fruits and
vegetables (e.g. berries, red cabbage, black rice). Several
different AC exist, differing in the number and substitution of
hydroxyl groups, the bonded sugar moieties, carboxylates type, and
bonding to sugars. A portion of the different classes of
anthocyanins includes: cyanidins, delphinidins, peonidins,
petunidins, and malvidins.
[0080] Current evidence indicates that parental AC are present
throughout the length of the gastrointestinal tract. At the colon,
AC's can be metabolized by microorganisms in the gut to several
metabolites. Thus, in theory, the small intestine and to a lesser
extent the large intestine can be exposed to significant amounts of
dietary parental AC. Despite the potential for benefits of dietary
AC on the gastrointestinal system, the particulars under conditions
of increased intestinal permeability, are mostly unknown. AC could
exert beneficial effects at the gastrointestinal tract through
direct and indirect effects. The indirect effects can be related to
the potential capacity of AC to modulate the microbiota, affecting
microbiota-mediated AC metabolism. Both a "healthier" microbiota
and the formation of select active metabolites could in turn
mediate AC indirect effects on intestinal health and lead to
systemic effects.
[0081] Reference is made hereinafter in detail to specific
embodiments of the invention. While the invention will be described
in conjunction with these specific embodiments, it will be
understood that it is not intended to limit the invention to such
specific embodiments. On the contrary, it is intended to cover
alternatives, modifications, and equivalents as may be included
within the spirit and scope of the invention as defined by the
appended claims. In the following description, numerous specific
details are set forth in order to provide a thorough understanding
of the present invention. The present invention may be practiced
without some or all of these specific details. In other instances,
well known process operations have not been described in detail, in
order not to unnecessarily obscure the present invention.
[0082] The present disclosure relates to compositions and methods
for improving intestinal health in a subject. In one example, an
intestinal health promoting composition is presented. The
composition can include a combination of cyanidins and delphinidins
(classes of anthocyanins), in an amount sufficient to treat
intestinal hyperpermeability. In one example, the cyanidins and the
delphinidins can be collectively present in an amount that
maintains tight junction integrity in the intestinal epithelial
cells. In another example, the cyanidins and the delphinidins can
be collectively present in an amount that restores tight junction
integrity in the intestinal epithelial cells.
[0083] Cyanidins and delphinidins can protect intestinal epithelial
cells against TNF.alpha. including loss of monolayer permeability
in transepithelial electrical resistance (TEER) and increased
paracellular transport of FITC-dextran. In contrast, malvidin,
peonidin, and petunidin do not seem to provide protective actions
to intestinal epithelial cells against TNF.alpha. induced TEER
permeability and do not seem to increase paracellular transport of
FITC-dextran. Without being bound by theory the protective effects
of the cyanidins and delphinidins, may be due to the presence of a
catechol group on the B-ring of the cyanidins and delphinidins. The
anthocyanins that the present inventors tested which did not
incorporate the B-ring did not exhibit these protective effects. In
one example, this activity can be selective. In another example,
the protective function can be dose dependent.
[0084] In another example, the present disclosure provides, a
method of treating a condition or disorder related to
gastrointestinal health in a subject comprising maximizing tight
junction integrity in epithelial cells of gastrointestinal tract of
the subject. In another example, a method of maximizing tight
junction integrity in epithelial cells of the gastrointestinal
tract of a subject is presented. In yet another example the present
disclosure provides, a method of treating intestinal
hyperpermeability. In some examples, these methods can be targeted
to (i) maintain and/or create a healthy microbiome in the
gastrointestinal system, (ii) maintain and/or create an
inflammatory balance within the gastrointestinal system, and/or
(iii) maintain and/or form intestinal cell barrier integrity.
[0085] In some embodiments, an intestinal health promoting
composition can include cyanidins, delphinidins, or a combination
thereof, and in some embodiments, these agents can be present in a
therapeutically effective amount. In one example, the composition
can include a combination of cyanidins and delphinidins in an
amount sufficient to treat intestinal hyperpermeability. In one
example, the cyanidins and the delphinidins can be collectively
present in an amount that maintains intestinal permeability. In
another example, the cyanidins and the delphinidins can be
collectively present in an amount that reduces intestinal
hyperpermeability. In yet another example, the cyanidins and
delphinidins can be capable of impacting the microbiome and can be
capable of providing anti-inflammatory properties.
[0086] The amount of cyanidins and delphinidins can vary in the
composition. In one example, the cyanidins and the delphinidins can
individually or collectively range from about 5 wt % to about 50 wt
% of the composition, or an active fraction of the composition. In
another example, the cyanidins and the delphinidins can
individually or collectively range from about 12 wt % to about 45
wt % of the composition, or an active fraction of the composition.
In a further example, the cyanidins and delphinidins can
individually or collectively range from about 12 wt % to about 25
wt % of the composition or an active fraction of the
composition.
[0087] The cyanidins and delphinidins can be derived from various
sources. In one example, a source of at least one of the cyanidins
and the delphinidins can be derived from a black rice component, a
blueberry component, a black current component, a crowberry
component, a bilberry component, a black chokeberry component, or a
combination thereof. In another example, the source of the
cyanidins and the delphinidins can be derived from the black rice
component, the blueberry component, and the black current
component. In yet another example, the source of the cyanidins and
the delphinidins can be derived from the black rice component and
the bilberry component. In a further example, the source of the
cyanidins and the delphinidins can be derived from the black
current component and the blueberry component. In another example,
the cyanidins and delphinidins can be artificially created or
synthesized (e.g. "synthetic").
[0088] In one example, the composition can include a black rice
component. The black rice component can be derived from black rice
kernels, black rice concentrate, black rice extract, black rice
powder, or a combination thereof. In one example, the black rice
component can be a black rice extract. For example, a liquid rice
extract can be obtained by concentrating black rice kernels and
passing the concentrated black rice kernels through a resin
absorption (chromatography column). In one example, the solvents
can be water and ethanol. In one example, the column can be eluted
with a 70 wt % ethanol and 30 wt % water solution. In another
example, the column can be eluted with 75 wt % ethanol and a 25 wt
% water solution. The eluent can then be concentrated into a
concentrated liquid extract, dried, and packaged. In another
example, the black rice extract can be derived from black rice
kernels. In one example, the black rice can be derived from Oryza
sativa L.
[0089] In one example, the black rice extract can make-up about 2.5
wt % to about 20 wt % of the composition, or an active fraction of
the composition. In another example, the black rice component can
be present from about 10 wt % to about 15 wt % of the composition,
or an active fraction of the composition. In a further example, the
black rice extract can be present from about 2.5 wt % to about 5 wt
% of the composition, or an active fraction of the composition. In
yet another example, the black rice component can be present from
about 2.5 wt % to about 7.5 wt % of the composition, or an active
fraction of the composition.
[0090] In one example, the black rice component can have
standardized anthocyanin content. In one example, the black rice
component can have a standardized anthocyanin content ranging from
about 10 wt % to about 30 wt %. In another example, the black rice
component can have a standardized anthocyanin content of about 20
wt %. In a further example, the black rice component can have a
standardized anthocyanin content of about 25 wt %. In one example,
the standard anthocyanin content can be measured by HPLC. In
another example, the standard anthocyanin content can be measured
by UV.
[0091] In another example, the composition can include the
blueberry component. The blueberry component can include a member
selected from the group consisting of blueberry fruit, blueberry
extract, blueberry concentrate, blueberry juice, blueberry powder,
or a combination thereof. In one example, the blueberry component
can be a blueberry powder. In another example, the blueberry
component can be a blueberry juice. In yet another example, the
blueberry component can be a blueberry powder derived from
blueberry juice. In a further example, the blueberry component can
be blueberry extract. For example, a blueberry fruit extract can be
obtained by extracting the blueberry fruit with water. The
extraction can be filtered and the filtrate can then be washed with
water and parsed with 75% ethanol thru resin adsorption. The
extract can then be concentrated under vacuum pressure and spray
dried. The concentrated extract was then grinded, sieved, and
packaged. In one example, the blueberry component can be a bog
blueberry component. In one example, the blueberry component can be
derived from Vaccinium uliginosum L.
[0092] In one example, the blueberry component can range from about
1 wt % to about 30 wt % of the composition or an active fraction of
the composition. In another example, the blueberry component can
range from about 1 wt % to about 10 wt % of the composition, or an
active fraction of the composition. In yet another example, the
blueberry component can range from about 25 wt % to about 30 wt %
of the composition, or an active fraction of the composition.
[0093] In one example, the blueberry component can have
standardized anthocyanin content. In one example, the blueberry
component can have a standardized anthocyanin content ranging from
about 0.5 wt % to about 30 wt %. In another example, the blueberry
component has a standardized anthocyanin content ranging from about
0.5 wt % to about 5 wt %. In yet another example, the blueberry
component can have a standardized anthocyanin content ranging from
about 20 wt % to about 30 wt %. In a further example, the blueberry
component can have a standardized anthocyanin content of about 25
wt %. In yet another example, the blueberry component can have a
standardized anthocyanin can be about 17% as measured by UV or
about 25% as measured by HPLC.
[0094] In one example, the composition can include the black
current component. In one example, the black current component can
include black current fruit, black current extract, black current
concentrate, black current juice, black current powder, or a
combination thereof. In another example, the black current
component can be a black current powder. In yet another example,
the black current component can be a black current juice. In a
further example, the black current component can be a black current
extract. In one example, the black current extract can be extracted
using water, a resin exchange, and ethanol. In one example, the
black current can be an ethylene oxide free product. In one
example, the black current component can be a derived from Ribes
nigrum.
[0095] In one example, the black current component can be from
about 0.5 wt % to about 15 wt % of the composition, or an active
fraction. In another example, the black current component can be
from about 1 wt % to about 5 wt % of the composition, or an active
fraction of the composition. In yet another example, the black
current component can be from about 10 wt % to about 15 wt % of the
composition, or an active fraction of the composition.
[0096] In one example, the black current component can have
standardized anthocyanin content. In one example, the black current
component can have a standardized anthocyanin content ranging from
about 20 wt % to about 40 wt %. In another example, the black
current component can have a standard anthocyanin content of about
30 wt %. In a further example, the black current component can have
a standardized anthocyanin content ranging from about 2.5 wt % to
about 10 wt %. In one example, the black current component can have
a standardized anthocyanin content of about 5 wt %.
[0097] In one example, the composition can include a crowberry
component. In one example, the crowberry component can be crowberry
fruit, a crowberry extract, a crowberry concentrate, a crowberry
juice, a crowberry powder, or a combination thereof. In another
example, the crowberry component can be crowberry fruit. In yet
another example, the crowberry component can be crowberry extract.
In one example, the crowberry extract can be extracted using water
and ethanol. In one example, the crowberry component can be derived
from Empetrum nigrum.
[0098] In one example, the crowberry component can range from about
1 wt % to about 30 wt % of the composition, or an active fraction
of the composition. In another example, the crowberry component can
range from about 5 wt % to 25 wt % of the composition, or an active
fraction of the composition. In a further example, the crowberry
component can range from about 5 wt % to about 15 wt % of the
composition, or an active fraction of the composition.
[0099] In one example, the crowberry component can have
standardized anthocyanin content. In one example, the crowberry
component can have a standard anthocyanin content ranging from
about 40 wt % to about 50 wt %. In a further example, the crowberry
component can have a standardized anthocyanin content of about 46.7
wt %.
[0100] In one example, the composition can include a bilberry
component. In one example, the bilberry component can be bilberry
fruit, bilberry extract, bilberry concentrate, bilberry juice,
bilberry powder, or a combination thereof. In one example, the
bilberry component can be a bilberry powder. In another example,
the bilberry component can be a bilberry extract. In yet another
example, the bilberry extract can be extracted with ethanol and
water. In one example, the ethanol to water can have an extract
ratio of 150:1. In one example, the bilberry component can be
derived from Vaccinium myrtillus.
[0101] The amount of the bilberry component in the composition can
vary. In one example, the bilberry component can range from about
0.5 wt % to about 30 wt % of the composition, or an active fraction
of the composition. In another example, the bilberry component can
range from about 2 wt % to about 20 wt % of the composition, or an
active fraction of the composition. In a further example, the
bilberry component can range from about 5 wt % to about 15 wt % of
the composition, or an active fraction of the composition.
[0102] In one example, the bilberry component can have standardized
anthocyanin content. In one example, the bilberry component can
have a standardized anthocyanin content ranging from about 1 wt %
to about 40 wt %. In another example, the bilberry component can
have a standardized anthocyanin content ranging from about 5 wt %
to about 25 wt %. In yet another example, the bilberry component
can have a standardized anthocyanin content ranging from about 20
wt % to about 40 wt %. In a further example, the bilberry component
can have an anthocyanin content of about 36 wt % as measured by
HPLC or about 25 wt % anthocyanins as measured by UV.
[0103] In one example, a source of at least one of the cyanidins
and the delphinidins can be derived from a black rice component, a
blueberry component, and a black current component. In another
example, the composition can include a black rice component, a
blueberry component, and a black current component at a ratio of
about 1:1:1. In another example, the black rice component, the
blueberry component, and the black current component can have a
ratio of about 1:1.4:4.3. In yet another example, the black rice
component, the blueberry component, and the black current component
can have a ratio of about 1:2:4.
[0104] In one example, the composition can further include a
prebiotic ingredient, or blend. Prebiotics can be naturally
occurring substances from fruit, vegetables, and grain. Prebiotics
can support the microbiome and gastrointestinal system by
stimulating growth or activity of at least 1 bacterium in the
colon.
[0105] In one example, the prebiotic, or prebiotic blend can
include inulin, fructooligosacharides, or a combination thereof. In
one example, the prebiotic, or prebiotic blend can include inulin.
In another example, the prebiotic, or prebiotic blend can include
fructooligosacharides. In a further example, the prebiotic, or
prebiotic blend can include inulin and fructooligosacharides. In
some examples, the prebiotic, or prebiotic blend can promote a
healthy microbiome by increasing the fuel available to the
beneficial bacteria.
[0106] In one example of the composition, the prebiotic, or
prebiotic blend can include inulin. In one example, the inulin can
be a powder. In one example, the inulin can be a chicory inulin. In
another example, the inulin can be a chicory inulin having
oligosaccharides and polysaccharides with fructose units linked by
.beta. (2-1) linkages. In another example, a source of the inulin
can be provided by Orafti.RTM. GR. Beneo-Orafti, Belgium. In some
examples, a source of the inulin can be derived from banana, onion,
flour, garlic, asparagus, wheat, rye, leeks, chicory root, sugar
beets, or a combination thereof. In one example, the inulin can be
recovered from the source by diffusion in hot water. In some
examples, the inulin can be hydrolyzed or partially hydrolyzed with
an enzymatic treatment.
[0107] In one example, the inulin can be present at various amounts
in the composition. In one example, the inulin can range from about
15 wt % to about 60 wt % of the composition, or an active fraction
of the composition. In another example, the inulin can range from
about 15 wt % to about 25 wt % of the composition, or an active
fraction of the composition. In yet another example, the inulin can
range from about 40 wt % to about 60 wt % of the composition, or an
active fraction of the composition.
[0108] In one example, the composition can include
fructooligosaccharides (FOS). In one example, the FOS can be short
chain FOS (having a degree of polymerization (DP) of .ltoreq.5).
Fructooligosaccharides can be derived from a variety of sources,
including grains, fruits, and vegetables. In one example, the short
chain FOS can be derived from sucrose. In another example, the
short chain FOS can be derived from sugarcane. In another example,
the short chain FOS can be derived from a non-GMO source. In yet
another example, the FOS can be galactooligosaccharides (GOS).
[0109] In one example, the FOS can be present in varying amounts in
the composition. In one example, the FOS can range from about 10 wt
% to about 40 wt % of the composition, or an active fraction of the
composition. In another example, the FOS can range from about 10 wt
% to about 20 wt % of the composition, or an active fraction of the
composition. In yet another example, the fructooligosaccharides can
range from about 25 wt % to about 40 wt % of the composition, or an
active fraction of the composition.
[0110] In one example, the composition can include inulin and
fructooligosacharides. When present, in one example, the inulin and
fructooligosacharides can collectively range from about 55 wt % to
about 95 wt % of the composition, or an active fraction of the
composition. In another example, the inulin and
fructooligosaccharides can collectively range from about 70 wt % to
about 90 wt % of the composition, or an active fraction of the
composition.
[0111] The ingredients above can be combined in a composition in
any variety of manners. Some, exemplary compositions are tabled
below. In the tables below, some of these examples include
excipients such as, silicon dioxide, while others display only the
active faction of the composition.
TABLE-US-00001 TABLE 1 Exemplary Formulation Ingredient Amount (mg)
Total wt % Black Rice Extract (25% AC) 770 12.28% Bilberry Extract
(36% AC) 122.22 1.95% Galactooligosacharides 2,769.23 44.16% Short
chain Fructooligosacharides 1,100 17.54% Inulin 1,500 23.92%
Silicon Dioxide 10 0.16% Total 6271.45 100.00%
TABLE-US-00002 TABLE 2 Exemplary Formulation Ingredient Amount (mg)
Total wt % Black Current Extract (5% AC) 1320 13.89% Blueberry
Juice Powder (0.8% AC) 2750 28.95% Galactooligosacharides 2,769.23
29.15% Short chain Fructooligosacharides 1,100 11.58% Inulin 1,500
15.79% Silicon Dioxide 60.769 0.64% Total 9499.999 100.00%
TABLE-US-00003 TABLE 3 Exemplary Formulation Ingredient Amount (mg)
Total wt % Blueberry Extract (24% AC) 144 3.63% Black Current
Extract (29% AC) 206 5.19% Black Rice Extract (19% AC) 618 15.57%
Short chain Fructooligosacharides 1,100 27.72% Inulin 1,900 47.88%
Total 3,968 100.00%
TABLE-US-00004 TABLE 4 Exemplary Formulation Ingredient Amount (mg)
Total wt % Black Rice Extract 120 4.12% Black Current Extract 60
2.06% Blueberry Extract 35 1.20% Fructooligosacharides 1,100 37.74%
Inulin 1,600 54.89% Total 2,915 100.00%
In some examples, the composition can be further formulated to
include additional excipients.
[0112] In one example, the composition can further include
epicatechins, catechins, or a combination thereof. In some
formulations, epicatechin and catechins can act as NADPH oxidase
inhibitors.
[0113] In one example, the composition can include a
pharmaceutically acceptable carrier. In another example, the
composition can include a sweetener, a preservative, a flavoring, a
thickener, or a combination thereof. In yet another example, the
composition can further include coatings, isotonic agents,
absorption delaying agents, binders, adhesives, lubricants,
disintegrants, coloring agents, flavoring agents, sweetening
agents, absorbents, detergents, emulsifying agents, antioxidants,
vitamins, minerals, proteins, fats, carbohydrates, or a combination
thereof. In some examples, the formulation can include polymers for
sustained release of a given compound. Nearly any number or type of
ingredients required in order to produce a specifically desired
composition or formulation can be used.
[0114] In one example, the composition can be in the form of an
oral dosage formulation. In one example, the oral dosage form
comprises a capsule, a tablet, a powder, a beverage, a syrup, a
gummy, a wafer, a confectionery, a suspension, or a food. In
another example, the oral dosage form can be in the form of a
capsule, a tablet, a soft gel, a lozenge, a sachet, a powder, a
beverage, a syrup, a suspension, or a food. In another example, the
oral dosage formulations can be formulated into a food or drink,
and provided, for example, as a snack bar, a cereal, a drink, a
gum, or in any other easily ingested form. In one example, the oral
dosage form can be incorporated into a liquid beverage such as
water, milk, juice, or soda. In another example, the oral dosage
form can be formulated into a nutritional beverage. The nutritional
beverage can be in a premixed formulation or can be a powdered mix
in that can be added to a beverage. In another example, the powder
mix in can be in the form of granules. In another example, the
composition can be a powder that can be sprinkled onto food.
[0115] In one example, the oral dosage form can be designed to be
administered to a subject in need thereof once per day. In one
example, the oral dosage form can be designed to be administered to
the subject in the morning. In another example, the oral dosage
form can be administered in the afternoon, or in the evening. In
yet another example, the dosage form can be designed to be
administered on an on and off and on again basis. For example, the
dosage form can be administered for 2 days on and 1 day off, 3 days
on with 2 days off, 3 days on with 4 days off, 4 days on with 3
days off, 5 days on with 2 days off; or 6 days on with 1 day off,
with each of these regimens repeating consecutively for a length of
time. In another example, the dosage regimen can alternate on and
off every day. The period of dosage can also vary. For example, the
dosage form can be designed to be administered for a two-weeks, a
three-weeks, a months, 6 weeks, two months, three months, four
months, 5 months, 6 months, a year, a year and a half, or
indefinitely.
[0116] The oral dosage form can include any of the compositions
identified herein. In one example, the oral dosage form comprises a
member selected from the group consisting of a black rice
component, a blueberry component, a black current component, a
crowberry component, a bilberry component, black chokeberry
component, or a combination thereof.
[0117] In one example, the oral dosage form can include the black
rice component and the black rice component can range from about
500 mg to about 800 mg of the oral dosage form. In another example,
the black rice component can have a standardized anthocyanin
content of about 15 wt % to about 30 wt %.
[0118] In one example, the oral dosage form can include the
blueberry component and the blueberry component can range from
about 100 mg to about 3,000 mg of the oral dosage form. In another
example, the blueberry component can range from about 50 mg to
about 500 mg. In a further example, the blueberry component can
range from about 2,000 mg to about 3,000 mg. In yet another
example, the blueberry component can have a standardized
anthocyanin content of about 0.5 wt % to about 25 wt %.
[0119] In one example, the oral dosage form can include the black
current component and the black current component can range from
about 200 mg to about 3,000 mg of the oral dosage form. In another
example, the black current component can range from about 50 mg to
about 500 mg. In a further example, the black current component can
range from about 2,000 mg to about 3,000 mg. In yet another
example, the black current component has a standardized anthocyanin
content of about 2.5 wt % to about 30 wt %.
[0120] In one example, the oral dosage form can include the
crowberry component and the crowberry component can range from
about 100 mg to about 1,000 mg of the oral dosage form. In one
example, the crowberry component has a standardized anthocyanin
content from about 1 wt % to about 50 wt %. In another example, the
crowberry component has a standardized anthocyanin content from
about 1 wt % to about 30 wt %. In yet another example, the
crowberry component has a standardized anthocyanin content ranging
from about 40 wt % to about 50 wt %.
[0121] In another example, the oral dosage form can include the
bilberry component and the bilberry component can range from about
100 mg to about 700 mg of the oral dosage form. In one example, the
bilberry component can have a standardized anthocyanin content of
about 30 wt % to about 40 wt %.
[0122] In a further example, the oral dosage form can include the
black chokeberry component and the black chokeberry component can
range from about 50 mg to about 700 mg. In another example, the
black chokeberry component can range from about 100 mg to about 600
mg. In yet another example, the black chokeberry component can
range from about 200 mg to about 500 mg. In one example, the black
chokeberry component can have a standardized anthocyanin content of
about 1 wt % to about 35 wt %
[0123] In one example, the oral dosage form can further include the
prebiotic, or prebiotic blend. The prebiotic, or prebiotic blend
can include inulin, fructooligosaccharides, or a combination
thereof. In one example, the inulin can be present in the oral
dosage in an amount of from about 1 grams to 2 grams. In another
example, the inulin can provide from about 1 gram to about 2 grams
of fiber in the oral dosage form.
[0124] In one example, the oral dosage form can include a
fructooligosaccharides (FOS). In one example, the FOS can range
from about 1 gram to about 1. 5 grams of the oral dosage form. In
another example, the FOS can range from about 3 grams to about 4
grams of the oral dosage form.
[0125] The oral dosage from can provide various amounts of
anthocyanins. In one example, the oral dosage from can collectively
provide from about 200 mg to about 300 mg of anthocyanins. In
another example, the oral dosage form can provide from 50 mg to 100
mg of anthocyanins. In one example, the oral dosage from can
provide about 80 mg of anthocyanins. In another example, the oral
dosage from can provide about 215 mg of anthocyanins.
[0126] The oral dosage form can also provide various amounts of
fiber. In one example, the oral dosage form can collectively
provide from about 1.5 grams to about 3 grams of fiber. In one
example, the oral form can provide about 2.6 g, 2.7 g, or 2.9 g, of
fiber.
[0127] Methods of making the present compositions are also
encompassed by the present invention. In one embodiment, the method
can include combining the ingredients, whether raw or extracted,
and processing the combined ingredients into the desired form of
the composition. In one example, the desired form of the
composition can be a tablet, capsule, powder, food, or beverage.
Individuals skilled in the art are of the procedures that can be
used to combine the components.
[0128] Further presented herein, are methods of treating intestinal
hyperpermeability in a subject. In one example, the method can
include administering an intestinal health promoting composition to
the subject. In one example, the intestinal health promoting
composition can be as described herein.
[0129] In another example, the intestinal health promoting
composition can be administered to the subject on a daily basis. In
one example, the administration can occur in the morning. In yet
another example, the administration can occur for an extended
period of time, such as, every day for at least 3 weeks, at least 4
weeks, at least 6 weeks, at least 8 weeks, at least 12 weeks, at
least 6 months, at least 9 months, at least a year, at least two
years, any period in-between these, or indefinitely.
[0130] Additionally presented, are methods of treating a condition
or disorder related to gastrointestinal health in a subject. In one
example, the method can include maximizing tight junction integrity
in epithelial cells of gastrointestinal tract of the subject. In
another example, the method can include reducing intestinal
hyperpermeability in the subject.
[0131] In one example, the method can improve the gastrointestinal
health of the subject. The improvement in gastrointestinal health
can vary.
[0132] In one example, the improvement can include an improvement
in the subject's bowel habits when compared to the subject's bowel
habits before administering the method. In another example, the
improvement in the subject's bowel habits can include a decrease in
straining during and after bowel movements. In yet another example,
the improvement can be a reduction in bloating, discomfort, gas, or
a combination thereof. In a further example, the improvement can be
reducing the gut permeability of the subject. In yet another
example, the improvement can be a reduction in the
symptoms/occurrence of leaky gut syndrome.
[0133] In a further example, the improvement in gastrointestinal
health can be a reduction in intestinal dysbosis. In one example,
the improvement can be evidenced by reduced calprotectin fecal
levels. In yet another example, the improvement can be an increase
short chain fatty acid levels.
[0134] In one example of the method, the condition or disorder can
include inflammation, inflammatory bowel disease, irritable bowel
syndrome, chronic intestinal diseases, celiac disease, Crohn's
disease, ulcerative colitis, food intolerances, dyspepsia, low
levels of chronic intestinal inflammation, gastrointestinal
infections, or a combination thereof.
[0135] In another example, the condition or disorder can be
inflammation and the inflammation can be reduced in the subject's
gastrointestinal tract when compared to the subject's
gastrointestinal inflammation before administering the method. In
another example, the reduction can be about a 50% reduction in
inflammation. In yet another example, the reduction can be about a
60% reduction in inflammation. In a further example, the reduction
can be about a 70% reduction in inflammation. In one example, the
reduction can be up to a 73% reduction in inflammation. In a
further example, supplementation for a 3 week period can result in
a modest reduction of inflammatory biomarkers.
[0136] In a further example, the condition or disorder can include
insufficient absorption of nutrients, endotoxemia, intestinal
hyperpermeability, or a combination thereof.
[0137] In another example, the condition or disorder can include
obesity, obesity associated pathologies, allergies, cardiovascular
conditions, type I diabetes, type II diabetes, rheumatoid
arthritis, insulin resistance, cancer, metabolic syndrome, asthma,
neurodegenerative diseases, or a combination thereof.
[0138] In one example, the condition or disorder can be a
cardiovascular condition or disorder. In one example, the
cardiovascular condition or disorder can be an increase in
high-density lipoprotein cholesterol for subjects that are not
taking cardiovascular medications. In another example, the
cardiovascular condition or disorder can be a decrease in HbA1c
levels. In yet another example, the decrease in HbA1c levels can be
from pre-diabetic levels to normal levels as measured by reducing
HbA1c levels from a range of 6-6.4% to below 6%. In a further
example, the decrease in HbA1c levels can be from diabetic levels
to pre-diabetic levels. This is a reduction from levels above 6.5%
to levels between 6% and 6.4%.
[0139] In a further example, the cardiovascular condition or
disorder can be related to an increase in plasma zonulin levels. In
one example, administering the method can decrease plasma zonulin
levels when compared to plasma zonulin levels prior to
administering the method. An increase in plasma zonulin can be
indicative of intestinal permeability. Zonulin can modulate
intestinal permeability by disassembling tight junctions and
allowing larger molecules, such as, lactulose to pass through.
Administering the compositions herein, can maximize tight junction
integrity; thereby, minimizing the amount of zonulin that passes
through the intestinal lining.
[0140] In another example, the condition can be peripheral insulin
resistance and the method can reduce peripheral insulin resistance.
In another example, the condition or disorder can be type I
diabetes or type II diabetes.
[0141] In yet another example, the condition or disorder can be a
nitric oxide related disorder, expression of iNOS, expression of
COX-2, NADPH oxidase, or a combination thereof. In one example, the
condition or disorder can stem from pathogens, antigens, and
pro-inflammatory factors that can pass through the tight
junctions.
[0142] In one example, the method can include maximizing tight
junction integrity or reducing intestinal hyperpermeability.
Maximizing tight junction integrity can include protecting the
gastrointestinal tract of the subject from TNF.alpha. induced
permealization of a monolayer of the epithelial cells. In another
example, the amount of the protecting can be concentration
dependent on an amount of cyanidins and delphinidins in the
subject's gastrointestinal tract. In one example, the amount of
protection is dose dependent.
[0143] In yet another example, the method can further include
increasing transepithelial electrical resistance in the epithelial
cells. In a further example, the method can include increasing FITC
dextran paracellular transport.
[0144] In one example, the condition or disorder can stem from
pro-inflammatory factors. In one example, the pro-inflammatory
factors can be advanced glycation end products. In another example,
the pro-inflammatory factors can be lipopolysaccharides. In a
further example the pro-inflammatory factors can include cytokines
tumor necrosis alpha (TNF-.alpha.), IL-6, or a combination
thereof.
[0145] In one example of the method, the condition or disorder can
relate to conditions or disorders associated with signaling
pathways NF-kB, ERK1/2, or a combination thereof.
[0146] In one example, maximizing of the tight junction integrity
can mitigate high fat induced intestinal permeabilization.
[0147] In another example, the epithelial cells can include Caco-2
cell monolayers.
[0148] In another example, the method can include optimizing a
balance of gut microbiota in the gastrointestinal tract. In one
example, optimizing the balance of gut microbiota can include
increasing commensal bacteria levels in the gastrointestinal tract
when compared to levels of commensal bacteria in the
gastrointestinal tract prior to administering the method. In
another example, the commensal bacteria can belong to
bifidobacteria genus. In one example, the commensal bacteria can
belong to bacteroidetes phylum. In another example, the commensal
bacteria can be bacterdies caccae, bacteriodes uniformis, or a
combination thereof. In yet another example, the increase in the
commensal bacteria after 8 weeks of daily administering the method
to the subject can be at least 20%. In another example, the
increase in commensal bacteria after 8 weeks of daily administering
the method to the subject can be about 5%, can about 10%, can be
about 15%, or can be about 25%.
[0149] In one example of the method, optimizing the balance of gut
microbiota can include increasing diversity of bacteria when
compared to the diversity of bacteria present in the gut prior to
administering the method. In one example, the diversity of bacteria
can include at least 200 strains. In another example, the method
can include decreasing harmful gut bacteria when compared to a
level of harmful gut bacteria present prior to administering the
method. In one example, the harmful gut bacteria can include
firmicutes. In some studies firmicutes have been found to make up a
higher portion of the gut microbiome in obese individuals. In yet
another example, the decreasing of the firmicutes after 8 weeks of
daily administering the method to the subject can be greater than a
15% reduction in firmicutes levels. In a further example, the
decreasing of the firmicutes after 8 weeks of daily administering
the method to the subject can be greater than a 5% reduction, about
a 10% reduction, about a 12% reduction, about a 15% reduction, or
about a 20% reduction.
[0150] In one example, the method can include optimizing the gut
microbiota by altering the firmicutes:bacteriodetes ratio. In one
example, the ratio of firmicutes:bacteriodetes can be decreased by
approximately 3% after 8 weeks of administering the method to the
subject. The firmicutes:bacteriodetes ratio can be a contributing
factor in obesity. Individuals with a high body mass index display
gut microbiota differences at the phylum level and can have high
firmicutes concentrations and can have low levels of bacteriodetes.
It is noted, that changes in the microbiota have not been
correlated to caloric content but rather to body mass index.
Accordingly, altering this ratio can allow for weight loss in the
individual if administered for a period of time.
[0151] In another example of the method for decreasing harmful gut
bacteria, the harmful gut bacteria can include Actinobacteria. In
one example, the reduction of the Actinobacteria after 8 weeks of
daily administering the method to the subject can be at least about
5%.
[0152] In yet another example of the method for decreasing harmful
gut bacteria, the harmful gut bacteria can include Helicobacter
pylori. Helicobacter pylori can be associated with ulcers and
heartburn. In a further example of the method, the harmful gut
bacteria can include Clostridium. In yet another example, the
harmful gut bacteria can include Klebisella.
[0153] In one example, the method can further include providing a
fuel source for commensal bacteria in the gut microbiome. In doing
this, the method can generate greater portions of commensal
bacteria which can ultimately lead to systemic health benefits.
[0154] In another example, a method of maximizing tight junction
integrity in epithelial cells of gastrointestinal tract of the
subject is provided. In one example, of this method the improvement
in tight junction integrity can include restoring tight junction
integrity. In another example of this method, the improvement can
include maintaining tight junction integrity. In a further example,
this method can provide systemic health benefits. These health
benefits can include improvements in conditions or disorders, such
as, celiac disease, IBS, Crohn's disease, ulcerative colitis, food
intolerances, allergies, dyspepsia, low levels of chronic
inflammation, obesity, type I diabetes, type II diabetes,
rheumatoid arthritis, insulin resistance, metabolic syndrome,
asthma, atopy, leaky intestinal barrier, tight junction barrier
dysfunction, plasma glucose levels, plasma free fatty acid levels,
reduced high density lipoprotein levels, hepatic steatosis,
firmicutes:bacteriodetes levels in the gut, abdominal bloating,
abdominal gas, abdominal pain, bowel function, growth of favorable
intestinal bacteria, short chain fatty acid production, plasma
zonulin levels, HbA1c levels, diabetes, prediabetes, nutrient
absorption, and combinations thereof.
[0155] The benefits of the various methods described above can be
achieved by administering a gastrointestinal health promoting
composition described above, to a subject. In one example, the
compositions can be administered on a daily basis for an extended
period of time. In another example, the compositions can be
administered on an on and off and on again basis based on a dosing
regimen. For example, the dosage regimen can be 2 days of
administration followed by 1 day without administration. In another
example, the dosage regimen could be 5 days of administration
followed by two days without administration. In yet another
example, the dosage regimen could be 3 days of administration
followed by 1, 2, 3, or 4 days without administration. In a further
example, the dosage regimen could be 4 days of administration
followed by 1, 2, 3, or 4 days without administration. In some
examples, the extended period of time can vary. In one example, the
extended period of time can be about 4 weeks. In another example,
the extended period of time can be about 6 weeks, about 8 weeks,
about 12 weeks, 16 weeks, 20 weeks, about 6 months, about 9 moths,
about 1 year, or a period of time greater than a year. In some
examples, the benefits of administering the method for a period of
time can increase with longer periods of administration.
Embodiments
[0156] In one embodiment presented herein, is an intestinal health
promoting composition, comprising a combination of cyanidins and
delphinidins, in an amount sufficient to treat intestinal
hyperpermeability.
[0157] In one embodiment of the composition, the cyanidins and the
delphinidins are collectively present in an amount that maintains
intestinal permeability.
[0158] In one embodiment of the composition, the cyanidins and the
delphinidins are collectively present in an amount that reduces
intestinal hyperpermeability.
[0159] In one embodiment of the composition, can include a source
of at least one of the cyanidins and the delphinidins that are
derived from a black rice component, a blueberry component, a black
current component, a crowberry component, a bilberry component,
black chokeberry component, or a combination thereof.
[0160] In one embodiment of the composition, the source of the
cyanidins and the delphinidins are derived from the black rice
component, the blueberry component, and the black current
component.
[0161] In one embodiment of the composition, the composition
comprises the black rice component and the black rice component is
derived from a member selected from the group consisting of black
rice kernels, black rice concentrate, black rice extract, black
rice powder, or a combination thereof.
[0162] In one embodiment of the composition, the black rice
component is a black rice extract.
[0163] In one embodiment of the composition, the black rice extract
is derived from black rice kernels.
[0164] In one embodiment of the composition, the black rice
component comprises from about 2.5 wt % to about 20 wt % of the
active fraction of the composition.
[0165] In one embodiment of the composition, the black rice
component comprises from about 10 wt % to about 15 wt % of the
active fraction of the composition.
[0166] In one embodiment of the composition, the black rice
component comprises from about 2.5 wt % to about 7.5 wt % of the
active fraction of the composition.
[0167] In one embodiment of the composition, the black rice
component has a standardized anthocyanin content ranging from about
10 wt % to about 30 wt %.
[0168] In one embodiment of the composition, the black rice
component has a standardized anthocyanin content of about 20 wt
%.
[0169] In one embodiment of the composition, the black rice
component has a standardized anthocyanin content of about 25 wt
%.
[0170] In one embodiment of the composition, the black rice
component is derived from Oryza sativa L.
[0171] In one embodiment of the composition, the composition
comprise the blueberry component and the blueberry component
comprises a member selected from the group consisting of blueberry
fruit, blueberry extract, blueberry concentrate, blueberry juice,
blueberry powder, or a combination thereof.
[0172] In one embodiment of the composition, the blueberry
component is a blueberry powder.
[0173] In one embodiment of the composition, the blueberry
component is a blueberry juice.
[0174] In one embodiment of the composition, the blueberry
component comprises from about 1 wt % to about 30 wt % of the
active fraction of the composition.
[0175] In one embodiment of the composition, the blueberry
component comprises from about 1 wt % to about 10 wt % of the
active fraction of the composition.
[0176] In one embodiment of the composition, the blueberry
component comprises from about 25 wt % to about 30 wt % of the
active fraction of the composition.
[0177] In one embodiment of the composition, the blueberry
component has a standardized anthocyanin content ranging from about
0.5 wt % to about 30 wt %.
[0178] In one embodiment of the composition, the blueberry
component has a standardized anthocyanin content ranging from about
0.5 wt % to about 5 wt %.
[0179] In one embodiment of the composition, the blueberry
component has a standardized anthocyanin content ranging from about
20 wt % to about 30 wt %.
[0180] In one embodiment of the composition, the blueberry
component is derived from Vaccinium uliginosum L.
[0181] In one embodiment of the composition, the composition
comprises the black current component and the black current
component comprises a member selected from the group consisting of
black current fruit, black current extract, black current
concentrate, black current juice, black current powder, or a
combination thereof.
[0182] In one embodiment of the composition, the black current
component is a black current extract.
[0183] In one embodiment of the composition, the black current
component comprises from about 0.5 wt % to about 15 wt % of the
active fraction.
[0184] In one embodiment of the composition, the black current
component comprises from about 1 wt % to about 5 wt % of the active
fraction.
[0185] In one embodiment of the composition, the black current
component has a standardized anthocyanin content ranging from about
20 wt % to about 40 wt %.
[0186] In one embodiment of the composition, the black current
component has a standard anthocyanin content of about 30 wt %.
[0187] In one embodiment of the composition, the black current
component is derived from Ribes nigrum.
[0188] In one embodiment of the composition, the composition
comprises the crowberry component and the crowberry component
comprises a member selected from the group consisting of crowberry
fruit, crowberry extract, crowberry concentrate, crowberry juice,
crowberry powder, or a combination thereof.
[0189] In one embodiment of the composition, the crowberry
component comprises crowberry fruit.
[0190] In one embodiment of the composition, the crowberry
component comprises crowberry extract.
[0191] In one embodiment of the composition, the crowberry
component comprises from about 1 wt % to about 30 wt % of the
active fraction of the composition.
[0192] In one embodiment of the composition, the crowberry
component comprises from about 5 wt % to about 25 wt % of the
composition.
[0193] In one embodiment of the composition, the crowberry
component has a standard anthocyanin content ranging from about 40
wt % to about 50 wt %.
[0194] In one embodiment of the composition, the crowberry
component has a standardized anthocyanin content of about 46.7 wt
%.
[0195] In one embodiment of the composition, the crowberry
component is derived from Empetrum nigrum.
[0196] In one embodiment of the composition, the composition
comprises the bilberry component and the bilberry component
comprises a member selected from the group consisting of bilberry
fruit, bilberry extract, bilberry concentrate, bilberry juice,
bilberry powder, or a combination thereof.
[0197] In one embodiment of the composition, the bilberry component
comprises bilberry extract.
[0198] In one embodiment of the composition, the bilberry component
ranges from about 0.5 wt % to about 30 wt % of the active fraction
of the composition.
[0199] In one embodiment of the composition, the bilberry component
ranges from about 2 wt % to about 20 wt % of the composition.
[0200] In one embodiment of the composition, the bilberry component
has a standardized anthocyanin content ranging from about 1 wt % to
about 30 wt %.
[0201] In one embodiment of the composition, the bilberry component
has a standardized anthocyanin content ranging from about 5 wt % to
about 15 wt %.
[0202] In one embodiment of the composition, the bilberry component
comprises 36 wt % anthocyanins as measured by HPLC or 25 wt %
anthocyanins as measured by UV.
[0203] In one embodiment of the composition, the bilberry component
is derived from Vaccinium myrtillus.
[0204] In one embodiment of the composition, the source of at least
one of the cyanidins and the delphinidins is derived from a black
rice component, a blueberry component, and a black current
component.
[0205] In one embodiment of the composition, the black rice
component, the blueberry component, and the black current component
have a ratio of about 1:1:1.
[0206] In one embodiment of the composition, the black rice
component, the blueberry component, and the black current component
have a ratio of about 1:1.4:4.3.
[0207] In one embodiment of the composition, the composition
further comprises a prebiotic blend.
[0208] In one embodiment of the composition, the prebiotic blend
comprises inulin.
[0209] In one embodiment of the composition, the inulin is a
chicory inulin having oligosaccharides and polysaccharides with
fructose units linked by .beta. (2-1) linkages.
[0210] In one embodiment of the composition, the inulin is derived
from banana, onion, flour, garlic, asparagus, wheat, rye, leeks,
chicory root, sugar beets, or a combination thereof.
[0211] In one embodiment of the composition, the inulin comprises
from about 15 wt % to about 60 wt % of the composition.
[0212] In one embodiment of the composition, the inulin comprises
from about 15 wt % to about 25 wt % of the composition.
[0213] In one embodiment of the composition, the inulin comprises
from about 40 wt % to about 60 wt % of the composition.
[0214] In one embodiment of the composition, the prebiotic blend
comprises fructooligosaccharides.
[0215] In one embodiment of the composition, the
fructooligosaccharides are short chain fructooligosaccharides
(DP.ltoreq.5).
[0216] In one embodiment of the composition, the short chain
fructooligosaccharides are derived from sucrose.
[0217] In one embodiment of the composition, the short chain
fructooligosaccharides are derived from sugarcane.
[0218] In one embodiment of the composition, the
fructooligosaccharides comprise from about 10 wt % to about 40 wt %
of the active fraction of the composition.
[0219] In one embodiment of the composition, the
fructooligosaccharides comprise from about 10 wt % to about 20 wt %
of the active fraction of the composition.
[0220] In one embodiment of the composition, the
fructooligosaccharides comprise from about 25 wt % to about 40 wt %
of the active fraction of the composition.
[0221] In one embodiment of the composition, the
fructooligosaccharides comprise galactooligosaccharides.
[0222] In one embodiment of the composition, the composition
further comprises a prebiotic blend of inulin and
fructooligosacharides.
[0223] In one embodiment of the composition, the inulin and
fructooligosacharides collectively comprise from about 55 wt % to
about 95 wt % of the composition.
[0224] In one embodiment of the composition, a combine source of
the cyanidins and the delphinidins comprise from about 5 wt % to
about 50 wt % of the composition.
[0225] In one embodiment of the composition, the composition
further comprises a pharmaceutically acceptable carrier.
[0226] In one embodiment of the composition, the composition
further comprises a sweetener, a preservative, a flavoring, a
thickener, or a combination thereof.
[0227] In one embodiment of the composition, the composition is an
oral dosage form.
[0228] In one embodiment of the composition, the oral dosage form
comprises a capsule, a tablet, a powder, a beverage, a syrup, a
gummy, a wafer, a confectionery, a suspension, or a food.
[0229] In one embodiment of the composition, the oral dosage form
comprises a powder.
[0230] In one embodiment of the composition, the oral dosage form
is designed to be administered to a subject in need thereof once
per day.
[0231] In one embodiment of the composition, the oral dosage form
is designed to be administered to the subject at morning.
[0232] In one embodiment of the composition, the oral dosage form
comprises a member selected from the group consisting of a black
rice component, a blueberry component, a black current component, a
crowberry component, a bilberry component, black chokeberry
component, or a combination thereof.
[0233] In one embodiment of the composition, the oral dosage form
comprises the black rice component and the black rice component
ranges from about 500 mg to about 800 mg of the oral dosage
form.
[0234] In one embodiment of the composition, the black rice
component has a standardized anthocyanin content of about 15 wt %
to about 30 wt %.
[0235] In one embodiment of the composition, the oral dosage form
comprises the blueberry component and the blueberry component
comprises from about 100 mg to about 3,000 mg of the oral dosage
form.
[0236] In one embodiment of the composition, the blueberry
component has a standardized anthocyanin content of about 0.5 wt %
to about 25 wt %.
[0237] In one embodiment of the composition, the oral dosage form
comprises the black current component and the black current
component comprises from about 200 mg to about 3,000 mg of the oral
dosage form.
[0238] In one embodiment of the composition, the black current
component has a standardized anthocyanin content of about 2.5 wt %
to about 30 wt %.
[0239] In one embodiment of the composition, the oral dosage form
comprises the crowberry component and the crowberry component
comprises from about 100 mg to about 1,000 mg of the oral dosage
form.
[0240] In one embodiment of the composition, the crowberry
component has a standardized anthocyanin content of about 1 wt % to
about 30 wt %.
[0241] In one embodiment of the composition, the oral dosage form
comprises the bilberry component and the bilberry component ranges
from about 100 mg to about 700 mg of the oral dosage form.
[0242] In one embodiment of the composition, the bilberry component
has a standardized anthocyanin content of about 30 wt % to about 40
wt %.
[0243] In one embodiment of the composition, the oral dosage form
comprises the black chokeberry component and the black chokeberry
component ranges from about 50 mg to about 700 mg of the oral
composition.
[0244] In one embodiment of the composition, the black chokeberry
component has a standardized anthocyanin content of about 1 wt % to
about 35 wt %.
[0245] In one embodiment of the composition, the oral dosage form
further comprises a prebiotic blend.
[0246] In one embodiment of the composition, the prebiotic blend
comprise from about 1 grams to 2 grams of the oral dosage form.
[0247] In one embodiment of the composition, the prebiotic blend
provides from about 1 gram to about 2 grams of fiber in the oral
dosage form.
[0248] In one embodiment of the composition, the oral dosage form
further comprises a fructooligosaccharide.
[0249] In one embodiment of the composition, the
fructooligosaccharide comprises from about 1 gram to about 1. 5
grams of the oral dosage form.
[0250] In one embodiment of the composition, the
fructooligosaccharide comprises from about 3 grams to about 4 grams
of the oral dosage form.
[0251] In one embodiment of the composition, the oral dosage form
comprises from about 200 mg to about 300 mg of anthocyanins.
[0252] Also presented herein in one embodiment, is a method of
treating intestinal hyperpermeability.
[0253] In one embodiment of the method, the method can include
administering an intestinal health promoting composition to a
subject.
[0254] In one embodiment of the method, the intestinal health
promoting composition comprises a combination of cyanidins and
delphinidins, in an amount sufficient to treat intestinal
hyperpermeability.
[0255] In one embodiment of the method, the intestinal health
promoting composition further comprises a prebiotic blend.
[0256] In one embodiment of the method, the intestinal health
promoting composition further comprises a
fructooligosaccharide.
[0257] In another embodiment of the method, the administering of
the intestinal health promoting composition can be on a daily
basis.
[0258] In one embodiment of the method, the administering can occur
in morning.
[0259] In one embodiment of the method, the administering can occur
for at least 3 weeks.
[0260] Further presented herein, in one embodiment, is a method of
treating a condition or disorder related to gastrointestinal health
in a subject comprising maximizing tight junction integrity in
epithelial cells of gastrointestinal tract of the subject.
[0261] In one embodiment of the method, the gastrointestinal health
of the subject is improved when compared to the health of the
gastrointestinal system in the subject prior to administering the
method.
[0262] In one embodiment of the method, the improved
gastrointestinal health of the subject comprises an improvement in
the subject's bowel habits when compared to the subject's bowel
habits prior to administering the method.
[0263] In one embodiment of the method, the improved
gastrointestinal health of the subject comprises reducing an
occurrence of bloating, discomfort, gas, or a combination thereof
in the subject when compared to the occurrence of bloating,
discomfort, gas, or the combination thereof prior to administering
the method.
[0264] In one embodiment of the method, the improved
gastrointestinal health of the subject comprises reducing
intestinal hyperpermeability.
[0265] In one embodiment of the method, the improved
gastrointestinal health of the subject comprises an improvement in
intestinal dysbiosis when compared to a level of intestinal
dysbiosis prior to administering the method.
[0266] In one embodiment of the method, the improved
gastrointestinal health of the subject comprises reduced
calprotectin fecal levels.
[0267] In one embodiment of the method, the improved
gastrointestinal health of the subject comprises an increase short
chain fatty acid levels.
[0268] In one embodiment of the method, the condition or disorder
comprises inflammation, inflammatory bowel disease, irritable bowel
syndrome, chronic intestinal diseases, celiac disease, Crohn's
disease, ulcerative colitis, food intolerances, dyspepsia, low
levels of chronic intestinal inflammation, gastrointestinal
infections, or a combination thereof.
[0269] In one embodiment of the method, the inflammation is reduced
overall.
[0270] In one embodiment of the method, the inflammation is reduced
by up to 73%.
[0271] In one embodiment of the method, supplementation for 3 weeks
with a gastrointestinal health promoting composition results in a
modest reduction in inflammatory biomarkers.
[0272] In one embodiment of the method, the condition or disorder
comprises insufficient absorption of nutrients, endotoxemia,
intestinal hyperpermeability, or a combination thereof.
[0273] In one embodiment of the method, the condition or disorder
comprises obesity, obesity associated pathologies, allergies,
cardiovascular conditions, type I diabetes, type II diabetes,
rheumatoid arthritis, insulin resistance, cancer, metabolic
syndrome, asthma, neurodegenerative diseases, or a combination
thereof.
[0274] In one embodiment of the method, the condition or disorder
is a cardiovascular condition.
[0275] In one embodiment of the method, the cardiovascular
condition is an increase in high-density lipoprotein cholesterol
for subjects that are not taking cardiovascular medications.
[0276] In one embodiment of the method, the cardiovascular
condition comprises a decrease in HbA1c levels.
[0277] In one embodiment of the method, the decrease in HbA1c
levels is from pre-diabetic levels to normal levels.
[0278] In one embodiment of the method, the cardiovascular
condition comprises a decrease in plasma zonulin levels.
[0279] In one embodiment of the method, the condition is peripheral
insulin resistance and the method reduces peripheral insulin
resistance.
[0280] In one embodiment of the method, condition or disorder is
type I diabetes or type II diabetes.
[0281] In one embodiment of the method, the condition or disorder
comprises a nitric oxide related disorder, expression of iNOS,
expression of COX-2, NADPH oxidase, or a combination thereof.
[0282] In one embodiment of the method, the condition or disorder
stems from pathogens, antigens, and pro-inflammatory factors that
pass through the tight junctions in the epithelial cells of the
gastrointestinal tract.
[0283] In one embodiment of the method, maximizing tight junction
integrity comprises protecting the gastrointestinal tract of the
subject from TNF.alpha. induced permeabilization of a monolayer of
the epithelial cells.
[0284] In one embodiment of the method, an amount of the protecting
is concentration dependent on an amount of cyanidins and
delphinidins in the subject's gastrointestinal tract.
[0285] In one embodiment of the method, the method further
comprises increasing transepithelial electrical resistance in the
epithelial cells.
[0286] In one embodiment of the method, the method further
comprises increasing FITC dextran paracellular transport.
[0287] In one embodiment of the method, the condition stems from
pro-inflammatory factors and the pro-inflammatory factors comprise
advanced glycation end products.
[0288] In one embodiment of the method, the condition stems from
pro-inflammatory factors and the pro-inflammatory factors comprise
lipopolysaccharides.
[0289] In one embodiment of the method, the condition stems from
pro-inflammatory factors and the pro-inflammatory factors comprise
cytokines tumor necrosis alpha (TNF-.alpha.), IL-6, or a
combination thereof.
[0290] In one embodiment of the method, the condition or disorder
relates to conditions or disorders associated with signaling
pathways NF-kB, ERK1/2, or a combination thereof.
[0291] In one embodiment of the method, the maximizing of the tight
junction integrity mitigates high fat induced intestinal
permeabilization.
[0292] In one embodiment of the method, the epithelial cells
comprise Caco-2 cell monolayers.
[0293] In one embodiment of the method, the method further
comprises optimizing a balance of gut microbiota in the
gastrointestinal tract.
[0294] In one embodiment of the method, optimizing the balance of
gut microbiota comprises increasing commensal bacteria levels in
the gastrointestinal tract above a level of the communsual bacteria
in the gastrointestinal tract prior to administering the
method.
[0295] In one embodiment of the method, the commensal bacteria
belong to bifidobacteria genus.
[0296] In one embodiment of the method, the commensal bacteria
belong to bacteroidetes phylum.
[0297] In one embodiment of the method, the commensal bacteria
comprise bacterdies caccae, bacteriodes uniformis, or a combination
thereof.
[0298] In one embodiment of the method, the increase in the
commensal bacteria after 8 weeks of daily administering the method
to the subject in need thereof was at least 20%.
[0299] In one embodiment of the method, optimizing the balance of
gut microbiota comprises increasing diversity of bacteria.
[0300] In one embodiment of the method, the diversity of bacteria
comprises at least 200 strains.
[0301] In one embodiment of the method, the method further
comprises decreasing a level harmful gut bacteria when compared to
the level of harmful gut bacteria prior to administering the
method.
[0302] In one embodiment of the method, the harmful gut bacteria
comprise firmicutes.
[0303] In one embodiment of the method, the decreasing of the
firmicutes after 8 weeks of daily administering the method to the
subject was greater than a 15% reduction.
[0304] In one embodiment of the method, a ratio of
firmicutes:bacteriodetes decreased approximately 3% after 8 weeks
of administering the method to the subject.
[0305] In one embodiment of the method, the harmful gut bacteria
comprise Actinobacteria.
[0306] In one embodiment of the method, the decreasing of the
Actinobacteria after 8 weeks of daily administering the method to
the subject was at least 5%.
[0307] In one embodiment of the method, the harmful gut bacteria
comprise Helicobacter pylori.
[0308] In one embodiment of the method, the harmful gut bacteria
comprise Clostridium.
[0309] In one embodiment of the method, the harmful gut bacteria
comprise Klebisella.
[0310] In one embodiment of the method, the method comprises
providing a fuel source for commensal bacteria.
EXAMPLES
Example 1--Bench Top Study of Anthocyanins (AC) Effect on Tumor
Necrosis Alpha-Induced Loss of Caco-2 Cell Barrier Integrity
[0311] The capacity of anthocyanins (AC) and seven AC-rich extracts
containing different types of AC were measured to determine their
ability to inhibit tumor necrosis alpha (TNF.alpha.) induced
permeabilization of Caco-2 cell monolayers. The AC were also tested
to determine the relationship between AC chemical
structure/conformation and to determine the extent of the AC
content to the extent of any protective capacity of the ACs to
inhibit TNF-.alpha. induced permeability of Caco-2 cell
monolayers.
Materials
[0312] Caco-2 cells were obtained from the American Type Culture
Collection (Rockville, Mass.).
[0313] Cell culture media and reagents, were from Invitrogen/Life
technologies (Grand Island, N.Y.).
[0314] HBSS 1.times. (21-022-CV) was obtained from Corning
(Manassas, Va.).
[0315] Millicell cell culture inserts 12 mm and 30 mm (0.4
.mu.m-pore polyester membranes) (PIHP01250 and PIHP03050,
respectively) were obtained from EMD Millipore (Hayward,
Calif.).
[0316] Fluorescein isothiocyanate (FITC)-dextran (46944-100MG-F)
and tumor necrosis factor-.alpha. human (TNF.alpha.) (T6674-10UG)
were obtained from Sigma Chem. Co. (St. Louis, Mo.).
[0317] Human interferon gamma (IFN-.gamma.) (#8901SC) was obtained
from Cell Signaling Technology (Danvers, Mass.).
[0318] The pure anthocyanins: delphinidin 3-O-glucoside (myrtillin)
(0938), cyanidin 3-O-glucoside (kuromanin) chloride (0915S), and
malvidin-3-O-glucoside (oenin) (0911S), were obtained from
Extrasynthese (Genay Cedex, France).
[0319] The anthocyanin rich powdered extracts were provided by
Pharmanex Research (Nu Skin Enterprises) and included: black
chokeberry extract powder (35% total AC), black rice extract (20%
total AC), wild blueberry extract (5% total AC), bilberry extract
(36% total AC), crowberry extract powder (30% total AC), blueberry
extract (25% total AC), and red grape extract (minimum 10% total
AC).
Methods
[0320] The resolution of chemical structures was performed by
molecular mechanics (MM2) according to Allinger using the routines
available in ChemBio3D Ultra 11.0.1 (Cambridge Scientific
Computing, Inc.). The structure of the anthocyanins appear in
FIGS.-5-9.
[0321] Caco-2 cells were cultured in a phenol red free minimum
essential medium (MEM) at 37.degree. C. and in a 5% (v/vl) CO.sub.2
atmosphere. The MEM media was supplemented with: 10% (v/v) fetal
bovine serum; antibiotics (50 .mu.g/ml penicillin, and 50 .mu.g/ml
streptomycin); 1% of non-essential amino acid (NEAA); and 1% of
sodium pyruvate. The cells were cultured for 21 days after
confluence to allow for differentiation into intestinal epithelial
cells. Over the course of the 21 days, the media was replaced every
3 days.
[0322] After 21 days, the caco-2 cells were differentiated into
polarized monolayers in Millicell cell inserts (30 mm, 0.4
.mu.m-pore polyester membranes) and were placed in 6-well plates.
The apical chamber consisted of 1.5 ml of media. After an initial
addition to the apical chamber of 15 .mu.l of a 1 mg/ml extract
solution dissolved in 20% (v/v) ethanol, cells were incubated at
37.degree. C. and 5% (v/vl) CO.sub.2.
[0323] Sampling occurred at time zero, 1 hour, and 3 hours. For the
initial time-point, 15 .mu.l of medium was withdrawn from the
apical chamber and the equivalent volume of extract was added. The
plate was gently agitated and a 200 .mu.l sample was immediately
withdrawn from the upper chamber. At 1 hour and 3 hours both the
apical and basolateral chambers were sampled. All samples were
immediately acidified with 2.5 .mu.L of 12M HCl and placed in a
80.degree. C. freezer until time of analysis.
[0324] Polyphenolic metabolites present after interacting with the
cell layer were identified in each extract by high performance
liquid chromatography (HPLC)-mass spectrometry (MS)/MS. The liquid
chromatography was performed on an Agilent series 1200 instrument
(Agilent Technologies, Santa Clara, Calif.) coupled with a
diode-array detector (DAD) that monitored the cells at wavelengths
280 and 520 nm. A Phenomenex Kinetex F5 pentafluorophenyl HPLC
column (2.6 .mu.M, 100.times.4.6 mm) with SecurityGuard.RTM.
cartridges (PFP, 4.0.times.2.0 mm) were used for the separation
with a flow rate of 0.70 mL min-1 and temperature of 37.degree. C.
Injector temperature was set to 4.degree. C. with a 7 .mu.L
injection. A binary gradient was employed consisting of 1.0% formic
acid (v/v) in water (mobile phase A) and 1.0% formic acid (v/v) in
acetonitrile (mobile phase B) (Fisher Scientific, Fair Lawn, N.J.).
The gradient was as follows: 1% B at 0 minutes, 7.5% B at 7 min,
7.6% B at 14 min, 10% B at 17 min, 12% B at 18.5 min, 30% B at 24
min, 90% B at 25 min, 1% B at 26 to 30 min. Mass spectral data was
acquired using an Agilent 6430 triple-quadrupole mass spectrometer
with electrospray injection (Agilent Technologies, Santa Clara,
Calif., USA) and multiple reaction monitoring (MRM) selected as the
mode of acquisition. Optimal MS/MS source parameters were set as
follows, nebulizer at 40 psi, capillary voltage +4000 V (or -3500
V), gas temperature 325.degree. C. and flow of 5 L-min. Sheath gas
was 250.degree. C. and sheath flow of 11 L-min.
[0325] Anthocyanin reference standards consisted of
malvidin-3-O-glucoside, cyanidine-3-O-glucoside,
cyanidine-3-O-galactoside, delphinidin-3-O-glucoside,
pelargonidin-3-O-glucoside, and peonidin-3-O-glucoside
(Extrasynthese, Genay Cedex, France). Anthocyanins that were
detected, but did not have a reference standard quantified by
equivalents of malvidin 3-O-glucoside. Phenolic acid reference
standards included syringic, vanillic, protocatechuic,
4-hydroxybenzoic, gallic (Sigma-Aldrich St. Louis, Mo.) and
3-O-methylgallic (Extrasynthese, Genay Cedex, France) acids.
Phloroglucinol aldehyde was supplied by Sigma-Aldrich (St. Louis,
Mo.).
[0326] To measure transepithelial electrical resistance (TEER)
cells were differentiated into polarized monolayers by culture in
transwell inserts (12 mm, 0.4 .mu.m-pore polyester membranes)
placed in 12-well plates. Epithelial cell monolayers, were
initially incubated for 24 hours with interpheron-.gamma. to
upregulate the TNF-.alpha. receptor. Then the monolayers in the
upper chamber were pre-incubated for 30 min with anthocyanin rich
extracts (1-10 .mu.g/ml) or purified compounds, or with myrtillin
chloride, kuromanin chloride, and oenin chloride at 0.25, 0.5, and
1 .mu.M concentration added to the apical compartment. TNF.alpha.
(5 ng/ml) was subsequently added to the basolateral compartment and
cells incubated for 6 more hours.
[0327] To determine TEER assessment, the incubation media was
removed from the apical and basolateral compartments, cells were
rinsed with HBSS 1.times. and the same solution was added to both
compartments and TEER was measured. TEER was measured using a
Millicell-ERS Resistance System (Millipore, Bedford, Mass.) that
includes a dual electrode volt-ohm-meter. see FIG. 10. TEER was
calculated as:
TEER=(R.sub.m-R.sub.1).times.A. (I)
wherein, R.sub.m, transmembrane resistance; R.sub.i, intrinsic
resistance of a cell-free media; and A, the surface area of the
membrane in cm.sup.2
[0328] To determine paracellular transport, apical-to-basolateral
clearance of FITC-dextran (4 kDa) was measured. After 6 hours of
incubation with TNF.alpha., the media was replaced in both
compartments with fresh serum and phenol red-free MEM; FITC-dextran
was then added to the apical compartment (100 .mu.M final
concentration) and allowed to incubate for 3.5 hours. Subsequently,
100 .mu.l of the medium in the basolateral compartment was
collected and diluted with 100 .mu.l of HBSS 1.times.. The
fluorescence was measured at .lamda.exc: 485 nm and .lamda.em: 530
nm in a fluorescence plate reader. Data was analyzed by one-way
analysis of variance (ANOVA) using Statview 5.0 (SAS Institute
Inc., Cary, N.C.). Fisher least significance difference test was
used to examine differences between group means. A P value <0.05
was considered statistically significant. Data is shown as
mean.+-.SEM.
Results
[0329] The AC concentration in the various extracts as assessed by
HPLC-MS/MS is shown in Table 5.
TABLE-US-00005 TABLE 5 Anthocyanin Content Black Black Wild Choke-
Kernel Blue- Crow- Blue- Red berry Rice berry Bilberry berry berry
Grape Anthocyanidin .mu.mol/g extract Cyanidin-3-O-galactoside 0.87
3.30 10.00 0.75 Cyanidin-3-O-glucoside 19.29 26.50 2.58 3.61
Cyanidin-3-arabinoside 9.89 2.86 3.41 17.26 Cyanidin-3-O-rutinoside
Delphinidin-3-O-galactoside 3.98 Delphinidin-3-O-glucoside 1.25
5.16 18.59 10.85 2.16 Delphinidin-3-arabinoside 0.55 4.25 4.25 0.00
Delphinidin-3-O-rutinoside 26.58 Petunidin-3-O-galactoside 0.90
1.95 Petunidin-3-O-glucoside 1.01 3.34 9.70 4.69
Petunidin-3-O-arabinoside 0.28 1.05 Petunidin-3-(6-acet) glucoside
1.30 Peonidin-3-O-glucoside 2.59 1.67 4.53 3.45
Malvidin-3-O-galactoside 1.44 3.55 2.53 5.07 Malvidin-O-glucoside
1.94 1.44 25.54 19.96 Malvidin-3-arabinoside 1.12 1.12 3.45
Malvidin-3-(6-acet) glucoside 1.11 Malvidin-3-(6-acet) galactoside
4.67 Malvidin-3-(6-coumaryl) 3.75 glucoside AC total content 30.05
29.09 8.48 36.24 82.00 58.31 46.92
Among the different extracts, total AC content varied between 8.5
and 82 .mu.mol/g per extract. For the individual ACs, the content
ranged between 0 to 30.05; 0 to 37.43; 0 to 9.70; 0 to 3.45; and 0
to 34.56 .mu.mol/g per extract for the different glycosides of
cyanidin, delphinidin, petunidin, peonidin and malvidin,
respectively.
[0330] The chemical structure and conformation of the
non-glycosylated forms (anthocyanidins) of the AC found in the
studied extracts are shown in FIGS. 5-9. While rings A and C are
identical for all the cyanidins, the positioning of the B ring with
respect to the C ring showed diedric angle values of 39, 37, 34,
39, and 43 degrees for cyanidin, delphinidin, petunidin, peonidin,
and malvidin respectively. The position of the B ring may play a
role in the beneficial properties of cyanidins and delphinidins. As
can be seen in FIGS. 7-9 petunidin, peonidin, and malvidin do not
incorporate the same positioning of the B ring.
[0331] The crowberry extract contained the highest total AC
content, 82 mol/g, and delivered one of the larger diversities of
individual ACs (cyanidin, delphinidin, petunidin, peonidin and
malvidin glycosides)(only bilberry AC were more diverse).
Accordingly, the crowberry extract was selected to determine the
concentration-dependent capacity of the AC-rich extracts to prevent
TNF.alpha.-induced permeabilization of Caco-2 monolayers was
assessed measuring both TEER and FITC-dextran paracellular
transport. Caco-2 monolayers were incubated in the presence of 5
ng/ml TNF.alpha. in the lower chamber (basolateral side of the
Caco-2 monolayer). This caused, a significant decrease (28%,
p<0.05) in TEER and an increase in FITC-dextran paracellular
transport (220%, p<0.05). These findings indicate that, under
the current experimental conditions, TNF.alpha. caused an increase
in permeabilization of the Caco-2 cell monolayer. The addition of
the crowberry extract (1-10 .mu.g/ml) to the upper chamber (apical
side of the Caco-2 monolayer) caused a concentration-dependent
recovery of the monolayer TEER and inhibition of TNF.alpha.-induced
FITC-dextran transport to the lower chamber. As can be seen in
FIGS. 11 and 12, TEER values increased as the amount of the extract
increased and paracellular transport values decreased as the amount
of extract increased. This indicated a dose dependent effect. Then
the relative capacity of all of the AC-rich extracts to inhibit
TNF.alpha.-induced Caco-2 cell monolayer permeabilization was
determined. At 5 .mu.g/ml concentrations, the extracts had
differential effects inhibiting TNF.alpha.-induced alterations on
TEER and FITC-dextran paracellular transport. See FIGS. 13 and
14.
TABLE-US-00006 TABLE 6 Extract Indicators Indicator
Extract/Formulation C Control - no extract; TNF.alpha. was not
induced TNF Tumor necrosis alpha induced 1 Black chokeberry extract
2 Black kernelled rice extract 3 Wild blueberry 4 Bilberry extract
5 Crowberry extract 6 Blueberry extract 7 Red grape
The black chokeberry (1), black kernel rice (2) and blueberry (6)
extracts were most effective at inhibiting TNF.alpha.-induced
alterations on TEER, whereas, the black kernel rice (2), bilberry
(4) and crowberry (5) extracts were most effective at inhibiting
TNF.alpha.-induced alterations on FITC-dextran paracellular
transport. The protective actions of the extracts on
TNF.alpha.-induced monolayer permeabilization was
concentration-dependent.
[0332] In order to evaluate if the AC present in the extracts could
be involved in the beneficial effects of the extracts on Caco-2
cell barrier integrity the correlation between TEER and the total
and individual extract AC content was assessed. TEER values were
not significantly correlated with the total AC content, or with the
content of peonidin, malvidin, and petunidin glycosides in the
extracts; however, TEER values were significantly (p<0.03)
correlated with the extracts' content of cyanidin (r: 0.73) and
delphinidin (r: 0.81) glycosides, see FIGS. 15 and 16, suggesting a
protective effect of these specific AC.
TABLE-US-00007 TABLE 7 Correlation between anthocyanin content and
protective capacity to prevent TNF.alpha. induced TEER decreases R
Anthocyanin (correlation coefficient) P Total Anthocyanins 0.100
0.48 Cyanidin 0.730 0.03* Delphinidin 0.810 0.03* Peonidin 0.270
0.48 Malvidin 0.001 0.99 Petunidin 0.140 0.48 *p < 0.05, only
cyanidin and delphinidin concentrations were significantly
positively correlated with protective capacity against TNFa induced
permeability, in other words, only cyanidins and delphinidins, not
total ACs or other individual ACs were protective.
TABLE-US-00008 TABLE 8 Epicatechin, Catechin, and Total Catechin
Content of the Extracts Extract/ Indicator Formulation Epicatechin
Catechin Total Catechins 1 Black chokeberry 8.4 4.7 13.1 extract 2
Black kernelled -- -- -- rice extract 3 Wild blueberry -- 0.9 0.9 4
Bilberry extract 2.9 2.7 5.6 5 Crowberry extract 9.5 5.7 15.2 6
Blueberry extract -- -- -- 7 Red grape 2.8 3.0 5.8
It was discovered that epicatechin and catechins prevents
TNF.alpha. induced losses of intestinal barrier integrity. See
FIGS. 17-20.
Example 2--Toxicity Study
[0333] A toxicity study was conducted to determine the toxicity of
a composition comprising cyanidins, delphinidins and a prebiotic
blend after 90 days of oral administration to Wistar rats. The
study also evaluated the dose response relationship and the
determination of the no-observed adverse effect level.
Materials--Environment
[0334] Healthy Wistar rats (Rattus norvegicus) were selected at
random to participate in the study. Then 60 female and 60 male
rates between 6-8 weeks old, were allocated to 6 different
groups.
[0335] Animals were individually housed in standard sized
(40.5.times.24.times.18.5 cm) polycarbonate cages with a sterilized
corn cob bedding. The bedding was changed weekly and more often as
necessary to keep the animals clean and dry.
[0336] The room temperature was maintained between 22.+-.3.degree.
C. with a relative humidity between 30-70%. Artificial light was
cycled for 12 hours of light and 12 hours of night. Air was changed
a minimum of 12 times during each hour in the animal room.
[0337] Animals were offered a pelleted rodent diet and filtered
water ad libitum. The water was placed in polycarbonate bottles
with stainless steel sipper tubes.
Methods
[0338] The selected rats were examined by a veterinarian and then
were acclimated to the test conditions for 5 days prior to the
initial dosing. The rats were allocated into six different groups
prior to initiation of the study using computer generated
randomization tables. The weight variation of the animals was
minimal and did not exceed .+-.20% of the mean weight.
TABLE-US-00009 TABLE 9 Mice and Diet Groupings Identification Dose
No. of Animals Group Treatment (mg/kg/day) Males Females G1 Control
- distilled water 0 01-10 61-70 G2 Low Dose 70 11-20 71-80 G3 Mild
Dose 350 21-30 81-90 G4 High Dose 700 31-40 91-100 G5 Recovery
Control 0 41-50 101-110 G6 Recovery High Dose 700 51-60 11-120
Oral formulations were prepared daily, on the day of dosing. The
formulation was dissolved in an aqueous solution. The high dose was
anticipated to be equivalent to a human dose.
[0339] The formulation consisted of blueberry extract (3.6%), black
currant extract (5.2%), black rice extract (15.6%), chicory inulin
(48%), and short chain fructooligosaccharides (27.6%) and was
administered orally once per day, at the same time each day, for 90
consecutive days. The quantity of the formulation administered was
10 ml/kg of body weight.
[0340] All animals were observed daily for clinical signs and
symptoms. Animals were observed individually in their home cage,
during removal/hand held observations, in an open field, and for
sensory reactivity. In the home cage, animals were observed for
posture, breathing rate and extent, clonic involuntary movement,
tonic involuntary movement, vocalisation, and palperbral closure.
During hand held observations animals were observed for palperbral
closure, lacrimation, eye and skin examination, piloerection, and
salivation. During field observation animals were observed for
gait, mobility, arousal, respiration, clonic movement, tonic
movement, vocalisation, rearing, urine pools, fecal boluses,
stereotype, and bizarre behavior. During sensory reactivity
observations, animals were assessed for sensory reactivity
including click response, touch response, tail pinch response, and
approach response.
[0341] The animals were weighed on day 1, weekly during the course
of the study, and on the day necropsy.
[0342] Urine was collected and analyzed for physical parameters and
microscopic examinations on the last week of treatment. In order to
collect the urine, animals were housed for 16-18 hours in metabolic
cages with graduated tubes attached to the bottom of the cages.
[0343] On day 91, blood samples were collected for hematology and
chemical analysis before necropsy and animals were fasted overnight
for 16-18 hours prior to the blood collection. On day 91, animals
in groups 1-4 were necropsied by an overdose of CO.sub.2 and
physically examined. The cranial, thoracic, and visceral cavities
were opened and examined macroscopically. The organs were trimmed
of tissues and fat and weighed. Organs and tissues were then
collected and preserved in 10% buffered formalin except for the
testes that were fixed in Modified Davidson's fluid. On day 105,
animals in groups 5-6 were necropsied, physically examined, and
their organs weighed.
Results
[0344] During the study, the physical observations were mostly
normal and/or consistent across all of the groups and determined to
be toxicology insignificant. No clonic or tonic behaviors were
observed. The only bizarre behavior was paper bitting. The paper
bitting was observed everyday in all of the groups, including the
control groups. The touch response and approach response were fast.
The tail pinch was observed as a flinch and the pupil response was
normal.
[0345] Hematology and clinical chemistry of the treated groups and
the control groups did not show any significant differences. Urine
chemistry also did not show any significant difference among all
the experimental animals and comparable to animals of the G1 and G4
groups.
[0346] Mean body weights were similar among the groups in the
study. The mean body weights gradually increased over the study. No
changes in food consumption were observed. Insignificant changes in
organ weight ratio were recorded and considered to be
toxicologically insignificant. Histopathological lesions were found
in both sexes and in the control groups with random distribution.
Accordingly, the lesions were considered to be random in
nature.
[0347] The study concluded that the formulation did not produce any
significant toxological changes in physical, physiological,
neurobehavioral, biochemical, hematological, and histopathological
parameters in any of the doses administered in the study. None of
the treatment related changes were considered to be of
toxicological significance.
Example 3--Animal Study 1
[0348] A mouse model of high fat diet induced obesity was used to
investigate the potential capacity of a diet rich in anthocyanins
to prevent and/or mitigate obesity induced intestinal inflammation,
increased intestinal barrier permeability, and insulin resistance.
The effects of anthocyanin supplementation on high fat diet induced
alterations in (a) intestinal inflammation, (b) intestinal
permeability, (c) gut microbiota, and (d) anthocyanin metabolism
were evaluated.
Materials
[0349] Sixty healthy male C57BL/6J mice (20-23 g) were obtained
from Jackson laboratories, and housed (4 mice/cage) in standard
stainless steel cages. An enrichment environment was provided with
the use of mouse houses and bedding. Mice were acclimated for one
week before starting the treatments. Mice were grouped and (10
mice/group/time point) fed one of: the control diet; the control
diet plus anthocyanins; a high fat diet; a high fat diet plus 2%
anthocyanins; a high fat diet plus 20% anthocyanins; or a high fat
diet plus 40% anthocyanins. The components in these diets are
described below.
[0350] The control diet, TD.06416, was obtained from Harlen Teklad,
WI. adjusted to approximately 10% calories from fat.
[0351] The high fat diet, TD.06414, was obtained from Harlan
Teklad, WI. and was a 60% fat diet. This diet was known to induce
weight gain and the development of obesity over time. In parallel
with weight gain, this diet was known to generate increases in
lipid levels (triglycerides, cholesterol, and adipocyte
accumulation); increases in blood glucose, the development of
insulin insensitivity; and when fed for extended periods of time,
diabetes.
[0352] The anthocyanin mix was obtained from Nu Skin Enterprises.
The mix was comprised of black rice extract, black current extract,
and blueberry extract.
Methods
[0353] Mice were housed in stainless steel cages. Four mice were
housed in each cage. 10 mice were placed in each group.
TABLE-US-00010 TABLE 10 Diet and Dosing Data Dose Diet mg
(AC)/kg/bodyweight Calories from fat Control (C) 0 10 wt % C +
anthocyanin (AC) 40 10 wt % High Fat (HF) 0 60 wt % HF + 2% AC 2 60
wt % HF + 20% AC 20 60 wt % HF + 40% AC 40 60 wt %
Each group of mice were fed one of the specialized diets above.
Food intake was monitored weekly. Body weight was gathered every
two weeks and a fresh diet prepared. Feces were gathered and tested
on weeks 0, 2, 4, 6, 8, 10, and 12. Urine was gathered on weeks 4
and 10. Gavage was collected on weeks 8 and 13. Blood was collected
on week 10 and midway between weeks 12 and 13. On week 14, the mice
were euthanized.
[0354] In order to measure intestinal permeabilization,
paracellular transport of FITC dextrin was given by gavage at 8
weeks on the diets.
[0355] Gene arrays were conducted in different sections of the
gastrointestinal tract.
[0356] Integrity of the tight junctions were measured as an
expression of tight junction proteins and the evaluation of
regulatory mechanisms.
[0357] Inflammation was determined by measuring F480+ macrophage
infiltration in the intestinal mucosa, CRP in plasma, TNF, MCP-1,
and iNOS expression in liver/intestinal mucosa.
[0358] In addition, ITT/GTTs were performed at weeks 10 and 11 to
determine the relationship between insulin sensitivity, intestinal
health, and the microbiota.
Results
[0359] During the course of the diet, each group of mice had steady
weight gain. The largest amount of weight gain was observed between
weeks 0-8 with smaller increases between weeks 8-12. Overall mice
on the high fat diet and the high fat diet+2% AC gained the most
weight over the study.
TABLE-US-00011 TABLE 12 Average Weight Gain Average weight gain (g)
Group Weeks 0-8 Weeks 8-12 Total Weight Gain Control (C) 10.2 -0.1
10.1 C + 40% 11.9 1.6 13.5 anthocyanin (AC) High Fat (HF) 16.43
2.87 19.3 HF + 2% AC 15.67 1.33 17 HF + 20% AC 15.6 2.6 18.2 HF +
40% AC 13.4 2.6 16
The mice fed diets supplemented with 60% of the calories from fat
sources, developed obesity, insulin resistance, and intestinal
permeabilization within 8-16 weeks of the diet.
[0360] Despite the weight gain, the amount of food consumed by mice
on the high fat diet was less than the amount of food consumed by
mice on the control diets. The amount of food ate by each of the
groups ranged between 2-5 grams each week during the study. Animals
on the control and the control+anthocyanin diet typically consumed
between 31/4 to 41/2 grams of food per week. Animals on the high
fat diet and high fat+diets typically consumed between 21/4 to 31/4
grams of food per week. Energy intake was similar among the groups.
This study solidified the link between diet and overall weight;
however, the study also showed that anthocyanin supplementation
generally had a lowering trend on overall weight gain.
[0361] In addition, the overall colon length and weight of the mice
was determined FIGS. 21, 22, and 23. FTC-dextran permeability,
intestinal permeability, and endotoxemia were also measured. FIGS.
24, 25, and 26, respectively. Mice with the high fat diet had the
most paracellular transport indicating higher intestinal
permeability. Supplementing the high fat diet with even small
amounts of anthocyanins, decreased or maintained lower levels of
intestinal permeability. Mice with a high fat diet also experienced
higher amounts of endotoxemia.
[0362] Blood was tested for glucose and insulin concentrations as
shown in FIGS. 27 and 28.
TABLE-US-00012 TABLE 13 Plasma Glucose and Insulin Concentrations
HF + HF + HF + C + Control HF 2% AC 20% AC 40% AC 40% AC Fasting
glucose 164 .+-. 8 230 .+-. 11 206 .+-. 17 225 .+-. 11 161 .+-. 1.3
200 .+-. 12 (mg/dl) Fasting Insulin 0.53 .+-. 0.04 1.22 .+-. 0.11
0.88 .+-. 0.16 1.08 .+-. 0.24 0.61 .+-. 0.10 0.47 .+-. 0.04 Fed
Glucose 124 .+-. 7 173 .+-. 18 150 .+-. 14 178 .+-. 15 136 .+-. 10
150 .+-. 13 (mg/dl) Fed Insulin 0.93 .+-. 0.15 1.60 .+-. 0.24 1.56
.+-. 0.30 1.50 .+-. 0.25 0.82 .+-. 0.17 0.99 .+-. 0.15
Mice on the high fat diet and on the diet comprising 20% AC
exhibited the highest glucose and insulin levels. In addition, as
the endotoxins increased so did the glucose tolerance and fasting
insulin levels. FIGS. 29 and 30. Endotoxin levels also increased as
IL-6 and IL-la levels increased; however, 1L-.beta. levels did not
show a trend that correlated to endotoxin levels. FIGS. 31-33.
[0363] HOMA-IR (homeostasis model assessment of insulin resistance,
this is a biomarker of insulin sensitivity), adiponectin, and
leptin levels are also shown in FIGS. 34, 35, and 36 respectively.
Adiponectin is an adipocyte-derived cytokine with anti-atherogenic,
anti-inflammatory, and anti-diabetic properties, which is decreased
in obesity. Leptin is another adipocyte-derived cytokine that plays
a role in the control of satiety and energy expenditure. Leptin
insensitivity, like insulin insensitivity has been associated with
weight gain and obesity. Ghrelin is a hormone known to stimulate
appetite.
[0364] In addition triglyceride and cholesterol levels were
measured in the plasma, liver, and the feces. See FIGS. 37-40. As
predicted, the high fat diet led to increased plasma triglycerides,
however, this increase was prevented by all three AC doses. FIG.
37
[0365] It is well established that a high fat diet leads to hepatic
steatosis (fatty liver) in this mouse, therefore, liver
triglyceride levels and cholesterol levels were also measured.
While triglycerides were significantly increased in the high fat
diet control group, the 40% AC diet prevented lipid accumulation in
the liver as triglyceride levels in the liver were similar in the
control group, control+AC and HF+40% AC groups. FIG. 38.
Cholesterol levels were also lower in all of the anthocyanin
containing diets than the high fat diet. FIGS. 39 and 40. Liver
triglyceride levels were also lower in the anthocyanin containing
diets than the high fat diet. FIG. 41. Representative images of
mice livers on each diet and their feces can be seen in FIGS. 42
and 43.
[0366] Interestingly, the highest triglyceride levels in the feces
were found in mice consuming the high fat plus 40% AC diet,
suggesting that the anthocyanins inhibited fat absorption, one
likely mechanism by which the AC blend may have prevented the high
fat diet induced hepatic steatosis and insulin insensitivity.
Plasma cholesterol levels were elevated in mice consuming the high
fat diet and the high fat+2% and 20% AC groups, only the 40% AC
diet attenuated plasma cholesterol levels. This may indicate that a
supplement having anthocyanin content can lower plasma total
cholesterol levels. All of the high fat diet groups exhibited
increased cholesterol levels in the feces, a parameter which did
not appear to be influenced by the addition of ACs to the diet.
Example 4--Human Study
[0367] This study was conducted to determine the effects of a
composition containing anthocyanins, inulin, and prebiotic fibers
on the microbial composition of obese adults.
Materials
[0368] A single dose of the composition, in Table 14 below,
contained 1.9 g inulin, 1.1 g fructooligosaccharides, 144 mg
blueberry, 206 mg black currant extract, and 618 mg black rice
extract.
TABLE-US-00013 TABLE 14 Composition Anthocyanin Active Ingredients
Weight Content Inulin (Beneo, Orafti GR) 1,900 mg N/A
Fructooligosaccharides 1,100 mg N/A (Ingredion, Nutraflora)
Blueberry Extract 144 mg 35 mg anthocyanins Black Currant Extract
206 mg 60 mg anthocyanins Black Rice Extract 618 mg 120 mg
anthocyanins
Methods
[0369] An initial screening was conducted two weeks prior to the
start of the study. During the initial screening potential
participants completed a review of their medical history,
indicating all concomitant therapies, and identifying any inclusion
and exclusion criteria. Potential participants were also measured
for resting blood pressure, heart rate, weight, height, and body
mass index. A pregnancy test was performed when applicable.
[0370] Fifty-one participants were enrolled in the study after
completing the 2-week trial period. The participants were
predominately female (73%) and Caucasian (93%). The accepted
participants were males and females between 20-60 years of age.
They had a BMI ranging from 29.9 to 39.9.+-.1 kg/m.sup.2 (29.2 to
40.6 kg/m.sup.2). Participants agreed to: maintain their level of
physical activity throughout the trial period; discontinue the use
of pre-biotic and pro-biotics and/or polyphenol supplements; and
discontinue eating foods containing anthocyanins (blueberries,
blackberries, cherries, grapes, grape juice, pomegranate,
raspberries, huckleberries, strawberries, and wine) for two prior
to the baseline assessment and during the study.
[0371] Participants were instructed to consume one sachet of the
formulation every morning with breakfast, by mixing it into their
beverage or food of choice. If participants forgot a dose they were
instructed to consume the next dose as soon as they remembered.
Participants did not consume more than one sachet per day.
Participants also recorded diaries for concomitant therapies,
adverse events, food records, daily bowel habits, and daily
abdominal discomfort. In addition, participants completed bloating
and flatulence questioners and had anthropometric measures
assessed.
[0372] The participants met with the investigational team at the
initial assessment and on days 0, 29, and 57 of the study. On day 0
and 57 stool samples, urine samples, and blood samples were
collected.
[0373] Stool samples were collected within 48 hours of day 0 and
day 57. The microbial composition and calprotectin were measured in
stool samples by a lab at U of Wisconsin. Microbial composition was
measured by 16s rRNA on the illumina platform and R&D Systems
(Minneapolis Mn.).
[0374] Laboratory parameters (CBC, electrolytes (N, K, Cl, Ca),
HbA1c, creatine, AST, ALT, GGT, and bilirubin) were assessed at the
initial assessment and at the end of the study.
[0375] Urine screenings were administered at the KGK Synergize
clinic. Blood parameters were measured by LifeLabs central
laboratory by a standard method.
TABLE-US-00014 TABLE 15 Tests Conducted Per Visit Protocol Pre
Screening Day 0 Day 29 Day 57 Height X Weight X X X X Heart Rate X
BP X X X Medical Review X X Physical Exam X X Pregnancy Test X
Fecal Samples X Formulation Dispensed X Diaries Dispensed/ X X X
Collected for Daily Recordings Bowel Habits Diaries and X
Questionnaires Dispensed Fecal Sample Kit X Dispensed Fecal Sample
Kit Collected X
[0376] Data entry and verification was executed according to KGK
Synergize's Standard Operating Procedures. Statistical analysis was
conducted on the results of all participants that consumed at least
80% of the treatment doses. Variables were tested for normality and
log-normality. Non-normal variables were analyzed by appropriate
non-parametric tests. Numerical efficacy and endpoints were
formally tested for significance by paired student t-tests.
Results
[0377] Forty-six of the 51 participants originally enrolled in the
study completed the study. Four of the participants dropped out
after the day 0 visit and before the first comparison measurement
on day 29. One individual dropped out between the visits on day 29
and day 57.
Microbial Diversity
[0378] Participants supplementing with the composition experienced
a favorable change in their microbial composition as demonstrated
by a change in their Firmicutes:Bacteriodetes ratio. FIG. 44. The
ratio decreased from 4.98 to 1.45 (Firmicutes decreased 74.9% to
59%; Bacteriodetes increased 13.8% to 34.5%). In addition,
Actinobacteria decreased from 8.5% to 3.4%. A total of 8 phyla (6
bacteria, 1 archaea, and 1 other) and 40 genera (7 Actinobacteria,
8 Bacteroidetes, 1 Eurychaeota, 21 Firmicutes, and 3
Proteobacteria) were changed following the supplementation.
TABLE-US-00015 TABLE 16 Phylum of Bacteria in Stool End Kingdom
Phylum Baseline of Study P-value Ratio Other Other 2.30E-03
3.63E-03 0.000 0.63 Archaea Euryarchaeota 1.12E-04 3.40E-05 0.010
3.29 Bacteria Other 6.52E-07 6.84E-07 1.000 0.95 Acidobacteria
1.56E-06 2.38E-05 0.268 0.07 Actinobacteria 8.54E-02 3.37E-02 0.000
2.54 Armatimonadetes 0.00E+00 4.46E-07 1.000 -- Bacteroidetes
1.38E-01 3.45E-01 0.000 0.40 Caldithrix 0.00E+00 1.15E-06 0.371 --
Chlamydiae 0.00E+00 1.41E-06 0.371 -- Chlorobi 0.00E+00 1.12E-06
1.000 -- Chloroflexi 1.32E-06 2.08E-05 0.168 0.06 Bacteria
Cyanobacteria 6.84E-04 5.88E-04 0.722 1.16 Firmicutes 7.49E-01
5.90E-01 0.000 1.27 Fusobacteria 5.15E-05 1.08E-04 0.990 0.47
Gemmatimonadetes 2.61E-07 1.89E-05 0.059 0.01 Lentisphaerae
4.28E-06 5.80E-05 0.017 0.07 NC10 2.83E-07 6.84E-07 0.789 0.41
Nitrospirae 2.45E-07 2.96E-06 0.201 0.08 OD1 0.00E+00 6.70E-07
1.000 -- Planctomycetes 0.00E+00 1.73E-05 0.014 -- Proteobacteria
1.43E-02 2.11E-02 0.093 0.68 SBR1093 4.65E-07 6.77E-06 0.295 0.07
Spirochaetes 5.23E-07 6.94E-07 1.000 0.75 Synergistetes 3.65E-05
4.24E-05 0.636 0.86 TM6 0.00E+00 1.34E-06 1.000 -- TM7 2.44E-04
1.03E-04 0.000 2.38 Tenericutes 4.50E-04 6.55E-04 0.030 0.69
Verrucomicrobia 9.38E-03 4.57E-03 0.433 2.05
TABLE-US-00016 TABLE 17 Proportion of Actinobacteria,
Bacteroidetes, and Firmicutes in Stool Actinobacteria Bacteroidetes
Firmicutes Mean .+-. SD (n) Mean .+-. SD (n) Mean .+-. SD (n)
Median (Min-Max) Median (Min-Max) Median (Min-Max) Within Group P
Value.sup..delta. Within Group P Value.sup..delta. Within Group P
Value.sup..delta. Proportion of Observed Species Baseline 0.085
.+-. 0.068 (45) 0.138 .+-. 0.096 (45) 0.749 .+-. 0.104 (45) (Day 0)
0.066 (0.002-0.349) 0.12 (0.006-0.473) 0.765 (0.492-0.936) End of
Study 0.034 .+-. 0.032 (45) 0.345 .+-. 0.159 (45) 0.590 .+-. 0.141
(45) (Day 57) 0.028 (0.004-0.15) 0.341 (0.003-0.639) 0.59
(0.287-0.89) Change from -0.052 .+-. 0.058 (45) 0.208 .+-. 0.176
(45) -0.159 .+-. 0.170 (45) Baseline to -0.038 (-0.199-0.029) 0.227
(-0.254-0.577) -0.18 (-0.501-0.226) End of Study p < 0.001* p
< 0.001* p < 0.001* Note: One participant was unable to
provide a fecal sample at the end of study. Max, maximum; Min,
minimum; n, number; SD, standard deviation. .sup..delta.Within
group comparisons were made using the paired Student t-test.
*Logarithmic transformation required to achieve normality
Probability values P .ltoreq. 0.05 are statistically
significant.
Inflammation
[0379] Following the study fecal calprotectin levels were reduced
indicating a trend for a reduction in gastrointestinal
inflammation. Calprotectin levels are tabulated below and
graphically presented in FIG. 45.
TABLE-US-00017 TABLE 18 Calprotectin Concentration Calprotectin
Concentration (pg/g) Mean .+-. SD (n) Median (Min-Max) Within Group
P Value.sup..delta. Baseline 1,024 .+-. 3,744 (43) (Day 0) 113
(7-23,026) End of Study 240 .+-. 375 (43) (Day 57) 106 (5-1,849)
Change from -784 .+-. 3,727 (43) Baseline to End of Studs -7
(-22,796-1,474) p = 0.107* Note: One participant was unable to
provide a fecal sample at the end of study. Two samples were not
available for calprotectin analysis. g, gram; Max, maximum; Min,
minimum; n, number; pg, picogram; SD, standard deviation.
.sup..delta.Within group comparisons were made using the paired
Student t-test. *Logarithmic transformation required to achieve
normality. Probability values P .ltoreq. 0.05 are statistically
significant.
Bowel Habits
[0380] A 5% increase was found on weeks 6 and 7 in the Bristol
stool score when compared to baseline. The number of bowel
movements did not change over the study; however, the bowel habits
were improved. FIG. 46. Participants reported a reduction in
straining before (33%) and while stopping defecation (51-54%) and a
reduction in incomplete bowel movements.
TABLE-US-00018 TABLE 19 Change in Daily Bowel Habits Proportion of
Bowel Number of Movements that Proportion of Bowel Bowel Proportion
of Bowel Had Movements that Bristol Stool Scale Movements Movements
that Had Straining to Stop Were Incomplete (n) (n/day) Straining to
Start (%) (%) Mean .+-. SD (n) Mean .+-. SD (n) (%) Mean .+-. SD
(n) Mean .+-. SD (n) Median (Min-Max) Median (Min-Max) Mean .+-. SD
(n) Median (Min-Max) Median (Min-Max) Within Group P Within Group P
Median (Min-Max) Within Group P Within Group P Value.sup..delta.
Value.sup..delta. Within Group P Value.sup..delta.
Value.sup..delta. Value.sup..delta. Bowel Habits Basline 3.88 .+-.
0.79 (46) 2.52 .+-. 1.09 (46) 17.7 .+-. 22.8 (46) 3.9 .+-. 10.3
(46) 22.9 .+-. 27.5 (46) (run-in) 3.87 (2.17-5.84) 2 (1-5) 6.7
(0-95.7) 0 (0-62.5) 11.4 (0-91.7) Change from -0.01 .+-. 0.59 (46)
-0.07 .+-. 0.88 (46) 1.5 .+-. 20.7 (46) -1.1 .+-. 8.2 (46) -3.8
.+-. 23.9 (46) Baseline to 0.04 (-1.4-1.21) 0 (-2-2) 0 (-40-45.7) 0
(-29.2-25) 0 (-68.3-80) Week 1 p = 0.924 p = 0.709* p = 0.420* p =
0.185* p = 0.191* Change from -0.00 .+-. 0.65 (46) -0.28 .+-. 0.89
(46) -1.1 .+-. 17.3 (46) 0.9 .+-. 9.6 (46) -8.7 .+-. 22.5 (46)
Baseline to -0.03 (-2.2-1.26) 0 (-2-1) 0 (-48-36.2) 0 (-16.7-41.7)
-4 (-79.2-41.7) Week 2 p = 0.994 p = 0.066* p = 0.083* p = 0.345* p
< 0.001* Change from 0.10 .+-. 0.55 (46) -0.17 .+-. 0.93 (46)
-2.5 .+-. 18.0 (46) 3.5 .+-. 16.1 (46) -8.7 .+-. 24.9 (46) Baseline
to 0.14 (-1.12-1.39) 0 (-2-2) 0 (-54-50) 0 (-10.7-75) -3.3 (-85-75)
Week 3 p = 0.225 p = 0.349* p = 0.089* p = 0.506* p < 0.001*
Change from 0.11 .+-. 0.70 (46) 0.04 .+-. 1.03 (46) -3.7 .+-. 19.1
(46) -1.2 .+-. 6.3 (46) -10.9 .+-. 21.9 (46) Baseline to 0.16
(-1.78-2.47) 0 (-2-2) -0.8 (-64.7-36.2) 0 (-25-25) -2.6 (-85-23.1)
Week 4 p = 0.309 p = 0.962* p = 0.004* p = 0.025* p < 0.001*
Change from 0.11 .+-. 0.54 (46) -0.26 .+-. 0.80 (46) -5.6 .+-. 18.0
(46) 0.5 .+-. 7.5 (46) -10.7 .+-. 27.2 (46) Baseline to 0.17
(-1.52-0.97) 0 (-2-1) -3.9 (-64.7-52.4) 0 (-10.7-37.5) -4.2
(-85-78.6) Week 5 p = 0.193 p = 0.056* p = 0.002* p = 0.124* p <
0.001* Change from 0.17 .+-. 0.58 (46) 0.02 .+-. 0.86 (46) -4.8
.+-. 14.1 (46) -1.2 .+-. 5.8 (46) -11.6 .+-. 20.6 (46) Basekline to
0.24 (-1.31-1.56) 0 (-2-3) -4 (-40-25) 0 (-25-14.3) -4.5 (-85-20)
Week 6 p = 0.051 p = 0.891* p < 0.001* p = 0.167* p < 0.001*
Change from 0.18 .+-. 0.53 (46) -0.15 .+-. 0.73 (46) -5.0 .+-. 16.9
(46) -2.0 .+-. 7.4 (46) -11.6 .+-. 24.2 (46) Baseline to 0.22
(-1.29-1.42) 0 (-1-2) 0 (-54.7-52.4) 0 (-33.9-14.3) -6.7
(-91.7-37.2) Week 7 p = 0.028 p = 0.284* p = 0.002* p = 0.046* p
< 0.001* Change from 0.18 .+-. 0.64 (46) -0.07 .+-. 0.95 (46)
-5.6 .+-. 19.1 (46) -2.2 .+-. 8.3 (46) -27 .+-. 108 (46) Baseline
to 0.28 (-1.39-1.71) 0 (-2-2) -1.7 (-47.6-50) 0 (-25-33.3) -6
(-721-50) Week 8 p = 0.063 p = 0.432* p < 0.001* p = 0.019* p
< 0.001* Max, maximum; Min, minimum; n, number; %, percent; SD,
standard deviation. .sup..delta.Within group comparisons were made
using the paired Student t-test. *Logarithmic transformation
required to achieve normality Probability values P .ltoreq. 0.05
are statistically significant.
Bloating, Discomfort, and Gas.
[0381] Participants experienced a reduction in bloating from weeks
3 through 8 when compared to baseline (week 4, 41%; week 5, 52%;
and week 8, 50%). FIG. 47.
TABLE-US-00019 TABLE 20 Bloating Proportion of Days Proportion of
Days With Bloating With Bloating Bloating Severity Due to Food Due
to Menstrual Cycle (n) (%) (%; Females Only) Mean .+-. SD (n) Mean
.+-. SD (n) Mean .+-. SD (n) Median (Min-Max) Median (Min-Max)
Median (Min-Max) Within Group P Value.sup..delta. Within Group P
Value.sup..delta. Within Group P Value.sup..delta. Bloating Results
Baseline 2.11 .+-. 2.07 (46) 26 .+-. 33 (46) 6.3 .+-. 10.9 (34)
(Day 0) 1.93 (0-9.07) 7 (0-100) 0 (0-35.7) Change from -0.10 .+-.
1.00 (46) -7.8 .+-. 23.6 (46) -2.1 .+-. 20.2 (34) Baseline to Week
1 -0.14 (-2.57-2.29) 0 (-64.3-42.9) 0 (-35.7-92.9) p = 0.125.lamda.
p = 0.034.dagger-dbl. p = 0.125.dagger-dbl. Change from -0.12 .+-.
1.15 (46) -7 .+-. 31 (46) -1.3 .+-. 22.5 (34) Baseline to Week 2
-0.14 (-2.36-3.07) 0 (-93-64) 0 (-35.7-92.9) p = 0.054.lamda. p =
0.167.dagger-dbl. p = 0.223.dagger-dbl. Change from -0.35 .+-. 1.21
(46) -8.7 .+-. 27.2 (46) -3.4 .+-. 13.9 (34) Baseline to Week 3
-0.24 (-3.71-2.14) -7.1 (-85.7-57.1) 0 (-35.7-42.9) p =
0.005.lamda. p = 0.056.dagger-dbl. p = 0.145.dagger-dbl. Change
from -0.61 .+-. 1.21 (46) -10.6 .+-. 27.5 (46) -4.2 .+-. 13.6 (34)
Baseline to Week 4 -0.21 (-3.71-1.43) -7.1 (-85.7-42.9) 0
(-35.7-28.6) p < 0.001.lamda. p = 0.024.dagger-dbl. p =
0.106.dagger-dbl. Change from -0.55 .+-. 1.34 (46) -13.4 .+-. 28.9
(46) -0.4 .+-. 21.1 (34) Baseline to Week 5 -0.14 (-3.71-3.07) -7.1
(-85.7-42.9) 0 (-35.7-85.7) p = 0.001.lamda. p = 0.005.dagger-dbl.
p = 0.462.dagger-dbl. Change from -0.59 .+-. 1.25 (46) -12 .+-. 33
(46) -2.5 .+-. 18.1 (34) Baseline to Week 6 -0.14 (-3.86-1.71) 0
(-100-43) 0 (-35.7-64.3) p < 0.001.lamda. p = 0.052.dagger-dbl.
p = 0.326.dagger-dbl. Change from -0.52 .+-. 1.24 (46) -10 .+-. 32
(46) -2.1 .+-. 19.3 (34) Baseline to Week 7 -0.14 (-3.5-1.93) -7
(-100-43) 0 (-35.7-92.9) p = 0.002.lamda. p = 0.101.dagger-dbl. p =
0.082.dagger-dbl. Change from -0.42 .+-. 1.01 (46) -13 .+-. 38 (46)
-2.9 .+-. 20.1 (34) Baseline to Week 8 -0.11 (-2.79-1.57) -7
(-100-79) 0 (-35.7-92.9) p < 0.001.lamda. p = 0.015.dagger-dbl.
p = 0.082.dagger-dbl. Max, maximum; Min, minimum; n, number; %,
percent; SD, standard deviation. .sup..delta.Within group
comparisons were made using the paired Student t-test.
.dagger-dbl.Within group comparisons were made using the
signed-rank test. .lamda.Square root transformation required to
achieve normality Probability values P .ltoreq. 0.05 are
statistically significant.
Participants also experienced a reduction in abdominal pain and
gas. Gas severity decreased overtime from 22% at week 5 to 11% at
week 8. FIG. 47.
TABLE-US-00020 TABLE 21 Change in Abdominal Pain Proportion of Days
Proportion of Days Abdominal Pain With Abdominal Pain With
Abdominal Pain Severity Due to Food Due to Menstrual Cycle (n) (%)
(%; Females Only) Mean .+-. SD (n) Mean .+-. SD (n) Mean .+-. SD
(n) Median (Min-Max) Median (Min-Max) Median (Min-Max) Within Group
P Value.sup..delta. Within Group P Value.sup..delta. Within Group P
Value.sup..delta. Abdominal Pain Results Baseline 1.90 .+-. 1.83
(46) 23.3 .+-. 29.6 (46) 5.5 .+-. 10.3 (34) (Day 0) 1.64 (0-8.43)
10.7 (0-100) 0 (0-35.7) Change from 0.09 .+-. 1.03 (46) -6.8 .+-.
20.3 (46) -0.8 .+-. 19.9 (34) Baseline to Week 1 0 (-1.95-3.07) 0
(-57.1-50) 0 (-35.7-92.9) p = 0.914* p = 0.025.dagger-dbl. p =
0.287.dagger-dbl. Change from 0.12 .+-. 1.15 (46) -4.0 .+-. 26.3
(46) 0.0 .+-. 23.2 (34) Baseline to Week 2 0 (-1.86-3.07) 0
(-85.7-64.3) 0 (-35.7-92.9) p = 0.972* p = 0.185.dagger-dbl. p =
0.305.dagger-dbl. Change from -0.18 .+-. 1.18 (46) -8.1 .+-. 20.1
(46) -1.7 .+-. 15.0 (34) Baseline to Week 3 -0.07 (-3.14-3.71) -7.1
(-71.4-28.6) 0 (-35.7-42.9) p = 0.050* p = 0.011.dagger-dbl. p =
0.448.dagger-dbl. Change from -0.44 .+-. 1.22 (46) -9.0 .+-. 23.3
(46) -1.7 .+-. 17.0 (34) Baseline to Week 4 -0.14 (-4.57-2.57) -7.1
(-78.6-42.9) 0 (-35.7-50) p = 0.006* p = 0.017.dagger-dbl. p =
0.555.dagger-dbl. Change from -0.35 .+-. 1.35 (46) -11.5 .+-. 24.8
(46) 1.3 .+-. 20.2 (34) Baseline to Week 5 -0.14 (-3.57-3.64) -7.1
(-85.7-42.9) 0 (-35.7-85.7) p = 0.037* p = 0.002.dagger-dbl. p =
0.723.dagger-dbl. Change from -0.32 .+-. 1.30 (46) -9.9 .+-. 28.9
(46) -1.3 .+-. 20.6 (34) Baseline to Week 6 -0.11 (-2.71-4.71) -3.6
(-100-42.9) 0 (-35.7-92.9) p = 0.031* p = 0.040.dagger-dbl. p =
0.288.dagger-dbl. Change from -0.33 .+-. 1.16 (46) -9.3 .+-. 26.5
(46) -0.4 .+-. 19.4 (34) Baseline to Week 7 -0.04 (-3.43-3.29) -7.1
(-100-42.9) 0 (-35.7-92.9) p = 0.014* p = 0.021.dagger-dbl. p =
0.304.dagger-dbl. Change from -0.28 .+-. 0.98 (46) -9 .+-. 34 (46)
-2.1 .+-. 19.9 (34) Baseline to Week 8 0 (-2.57-2) -7 (-100-79) 0
(-35.7-92.9) p = 0.014* p = 0.051.dagger-dbl. p = 0.118.dagger-dbl.
Max, maximum; Min, minimum; n, number; %, percent; SD, standard
deviation. .sup..delta.Within group comparisons were made using the
paired Student t-test. .dagger-dbl.Within group comparisons were
made using the signed-rank test. .lamda. Square root transformation
required to achieve normality Probability values P .ltoreq. 0.05
are statistically significant.
TABLE-US-00021 TABLE 22 Change in Gas Proportion of Days Proportion
of Days With Gas Gas Severity With Gas Due to Menstrual Cycle (n)
Due to Food (%) (%; Females Only) Mean .+-. SD (n) Mean .+-. SD (n)
Mean .+-. SD (n) Median (Min-Max) Median (Min-Max) Median (Min-Max)
Within Group P Value.sup..delta. Within Group P Value.sup..delta.
Within Group P Value.sup..delta. Gas Results Baseline 2.40 .+-.
1.96 (46) 36 .+-. 36 (46) 5.3 .+-. 10.3 (34) 2.04 (0-8.5) 18
(0-100) 0 (0-35.7) Change from -0.02 .+-. 1.09 (46) -10.7 .+-. 29.6
(46) -1.5 .+-. 19.5 (34) Baseline to Week 1 0.07 (-2.57-3.36) -7.1
(-71.4-92.9) 0 (-35.7-92.9) p = 0.843* p = 0.003.dagger-dbl. p =
0.136.dagger-dbl. Change from -0.02 .+-. 1.16 (46) -10 .+-. 36 (46)
-1.5 .+-. 18.2 (34) Baseline to Week 2 -0.11 (-2.71-1.93) -7
(-86-79) 0 (-35.7-78.6) p = 0.462* p = 0.031.dagger-dbl. p =
0.210.dagger-dbl. Change from -0.16 .+-. 1.29 (46) -12.6 .+-. 27.3
(46) -2.3 .+-. 13.6 (34) Baseline to Week 3 -0.25 (-2.5-4.57) -7.1
(-85.7-50) 0 (-35.7-42.9) p = 0.340* p = 0.004.dagger-dbl. p =
0.325.dagger-dbl. Change from -0.36 .+-. 1.11 (46) -9 .+-. 35 (46)
-3.2 .+-. 14.3 (34) Baseline to Week 4 -0.21 (-4-2.36) -7 (-86-93)
0 (-35.7-42.9) p = 0.062* p = 0.118.dagger-dbl. p =
0.156.dagger-dbl. Change from -0.52 .+-. 1.31 (46) -17.2 .+-. 29.8
(46) 1.1 .+-. 20.0 (34) Baseline to Week 5 -0.29 (-4.14-2.07) -7.1
(-85.7-35.7) 0 (-35.7-85.7) p = 0.007* p < 0.001.dagger-dbl. p =
0.608.dagger-dbl. Change from -0.35 .+-. 1.38 (46) -16 .+-. 32 (46)
-1.5 .+-. 17.8 (34) Baseline to Week 6 -0.36 (-4-2.86) -7 (-100-36)
0 (-35.7-64.3) p = 0.056* p = 0.007.dagger-dbl. p =
0.408.dagger-dbl. Change from -0.34 .+-. 1.22 (46) -14 .+-. 34 (46)
-1.5 .+-. 16.9 (34) Baseline to Week 7 -0.25 (-3.71-2.5) -7
(-100-43) 0 (-35.7-78.6) p = 0.043* p = 0.020.dagger-dbl. p =
0.166.dagger-dbl. Change from -0.26 .+-. 1.07 (46) -16 .+-. 39 (46)
-1.5 .+-. 20.4 (34) Baseline to Week 8 -0.21 (-2.57-2.5) -7
(-100-79) 0 (-35.7-92.9) p = 0.035* p = 0.006.dagger-dbl. p =
0.212.dagger-dbl. Max, maximum; Min, minimum; n, number; %,
percent; SD, standard deviation. .sup..delta.Within group
comparisons were made using the paired Student t-test.
.dagger-dbl.Within group comparisons were made using the
signed-rank test. .lamda. Square root transformation required to
achieve normality Probability values P .ltoreq. 0.05 are
statistically significant.
Vital Signs
[0382] Participants experienced a clinically insignificant
reduction in diastolic blood pressure from baseline to the end of
the study.
TABLE-US-00022 TABLE 23 Vital Signs Systolic Blood Diastolic Blood
Heart Rate Pressure (mmHg) Pressure (mmHg) (BPM) Weight (kg) BMI
(kg/m.sup.2) Mean .+-. SD (n) Mean .+-. SD (n) Mean .+-. SD (n)
Mean .+-. SD (n) Mean .+-. SD (n) Median (Min-Max) Median (Min-Max)
Median (Min-Max) Median (Min-Max) Median (Min-Max) Within Group P
Within Group P Within Group P Within Group P Within Group P
Value.sup..delta. Value.sup..delta. Value.sup..delta.
Value.sup..delta. Value.sup..delta. Vitals and Anthropometric
Measures Screening 119.8 .+-. 11.8 (51) 76.5 .+-. 10.1 (51) 74.1
.+-. 12.0 (51) 95.7 .+-. 14.2 (51) 34.2 .+-. 3.2 (51) 119.3
(92.3-146) 75 (57-98.7) 75 (55-109) 95 (71.6-137.1) 33.7
(29.2-40.6) Baseline 120.0 .+-. 12.0 (51) 79.4 .+-. 9.2 (51) 72.9
.+-. 10.4 (51) 95.2 .+-. 14.0 (51) 34.1 .+-. 3.1 (51) (Day 0) 121.7
(95.7-143) 80.3 (58-101.7) 74 (46-91) 94 (71.4-134.4) 33.6
(28.7-40.8) Mid-point 120.5 .+-. 12.5 (50) 77.3 .+-. 8.7 (50) 74.0
.+-. 10.0 (50) 94.7 .+-. 12.9 (50) 34.1 .+-. 3.1 (50) (Day 29)
122.3 (92.3-151) 77.8 (56-102) 72.5 (52-100) 94.8 (71.6-128.3) 33.9
(28.5-40.8) End of 120.3 .+-. 11.1 (47) 76.7 .+-. 8.3 (47) 71.6
.+-. 10.2 (47) 94.4 .+-. 13.1 (47) 33.9 .+-. 3.2 (47) Study 120
(98.7-141) 75.7 (60.3-96.3) 70 (49-97) 94 (71-127.4) 33.5 (28.7-41)
(Day 57) Change 0.7 .+-. 11.6 (50) -1.9 .+-. 8.6 (50) 1.2 .+-. 9.3
(50) 0.36 .+-. 1.38 (50) 0.14 .+-. 0.51 (50) from 0.5 (-28.7-35)
-0.8 (-23.7-16.7) 1 (-19-34) 0.25 (-2.3-4.6) 0.11 (-0.95-1.8)
Baseline to p = 0.663 p = 0.122 p = 0.365 p = 0.071 p = 0.065
Mid-point Change 0.4 .+-. 10.0 (47) -2.9 .+-. 8.2 (47) -0.9 .+-.
8.7 (47) 0.06 .+-. 2.02 (47) 0.04 .+-. 0.73 (47) from 2.3
(-25.7-22.7) -3.7 (-18.7-17.7) -1 (-26-20) 0.1 (-5.2-5.4) 0.04
(-1.72 -2.11) Baseline to p = 0.799 p = 0.020 p = 0.506 p = 0.847 p
= 0.716 End of Study BPM, beats per minute; kg, kilograms; Max,
maximum; mmHg, m, meter; millimeters of mercury; Min, minimum; N,
number; SD, standard deviation. .sup..delta.Within group
comparisons were made using the paired Student t-test. Probability
values P .ltoreq. 0.05 are statistically significant
CONCLUSION
[0383] Overall the study indicated that the formulation was capable
of improving the intestinal microbiome in mildly to moderately
obese yet otherwise healthy individuals. Over the study,
participants experienced a significant reduction in the Firmicutes
and a significant increase in Bacteroidetes proportions. The
Firmicutes:Bacteroidetes ratio decreased from day 0 to day 57 with
supplementation. The ratio changed from 4.98 to 1.45. In addition,
Actinobacteria levels decreased. These three phyla make up about
97% of the bacterial composition at baseline (97.2%) and following
supplementation (96.9%).
[0384] Thirty adverse events were recorded by 18 participants in
the study. Of the thirty reported incidents, eleven were assessed
as unlikely and 7 were assessed as unrelated. Two of the adverse
events were determined to be likely and included feces
discoloration and tooth discoloration. The ten possible adverse
events included reports of abdominal discomfort, diarrhea, feces
discoloration, frequent bowel movements, and vomiting.
[0385] Formulations, methods for the production of these
formulations, and uses for the formulations have been described. It
will be readily apparent to those skilled in the art, however
various changes and modifications of an obvious nature may be made
without departing from the spirit of the invention, and all such
changes and modifications are considered to fall within the scope
of the invention as defined by the appended claims. Such changes
and modifications would include, but not be limited to, the
incipient ingredients added to affect the capsule, tablet, powder,
lotion, food, powder, or bar manufacturing process as well as
vitamins, flavorings, and carriers. Other such changes or
modifications would include the use of herbs or other botanical
products containing the combinations of the preferred embodiments
disclosed above. Many additional modifications and variations of
the embodiments described herein may be made without departing from
the scope, as is apparent to those skilled in the art. The specific
embodiments described herein are offered by way of example
only.
* * * * *